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<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
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<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
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<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
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<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
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'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li><span style="font-weight: 400;">An innovative technology with template switching and UMIs</span></li>
<li><span style="font-weight: 400;">Ultra-low input capability, down to 50 pg for total RNAs</span></li>
<li><span style="font-weight: 400;">Capture the widest possible diversity of RNAs for rich content</span></li>
<li><span>Get high sensitivity data even from difficult samples, such as degraded, FFPE samples</span></li>
<li><span style="font-weight: 400;">Enjoy a fast, easy, single tube protocol</span></li>
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<p></p>
<p></p>
<h4></h4>
<center><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/dplex-total-manual.pdf" target="_blank" title="dplex total rnaseq user manual"><strong>Download the manual</strong></a></center>
<p></p>
<p>D-Plex Total RNA-seq Library Preparation Kit is a tool designed for the study of the whole coding and non-coding transcriptome. <span>The kit is using the</span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank"><span> </span>D-Plex technology</a><span><span> </span>to generate directional libraries for Illumina sequencing directly from total RNAs, mRNAs that has already been enriched by poly(A) selection, or RNAs that has already been depleted of rRNAs.</span></p>
<p><span>The D-Plex technology utilizes two innovative ligation-free mechanisms - poly(A) tailing and template switching - to produce sequencing libraries from ultra-low input amounts, down to 50 pg for total RNAs, mRNAs or rRNA-depleted RNAs. Combined with unique molecular identifiers (UMI), this complete solution delivers a high-sensitivity detection method for comprehensive analysis of the transcriptome. It accurately measures gene and transcript abundance and captures the widest possible diversity of RNAs, including known and novel features in coding and non-coding RNAs.</span></p>
<p><span></span><span>D-Plex Total RNA-seq Kit offers a time saving protocol that can be completed within 5 hours and requires minimal hands-on time. <span>The library preparation takes place in a </span><span>single tube</span><span>, increasing the efficiency tremendously. This new solution has been extensively validated for both intact and highly degraded RNA samples, including that derived from FFPE preparations.</span></span></p>
<p><span><span>D-Plex Total RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Unique Dual Indexes were designed and validated to fit the D-Plex technology and are available separately:</span></span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C </a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<div class="extra-spaced" align="center"></div>
<div class="row">
<div class="carrousel" style="background-position: center;">
<div class="slick">
<div>
<h3>Greater sensitivity to detect novel transcripts</h3>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/CPM_totalrna.png" alt="total RNA kit" /></center></div>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq kit enables great transcript detection, even when starting from very low RNA inputs or working with challenging FFPE samples. Increased number of transcripts detected at 1x coverage is an indicator of greater sensitivity.</p>
</div>
</div>
<div>
<h3>Highest diversity to facilitate biomarker discovery</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_rdep_biotypes.png" alt="total RNA diversity" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq libraries capture efficiently all RNA biotypes present in the sample of interest, both coding and non-coding RNAs or small and long RNAs.</p>
</div>
</div>
<div>
<h3>Consistent expression profiling across a wide range of RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_and_umeg_rdep_violin_tpm_10.png" alt="total RNA" width="450px" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Gene expression profiling (including protein coding exons and introns) obtained with D-Plex Total RNA-seq kit is conserved for decreasing RNA inputs, keeping transcript diversity across the range of RNA inputs.</p>
</div>
</div>
<div>
<h3>Superior library complexity at low RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/robustness_corr_HUR_HuMEg_rrna.png" alt="total RNA" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Correlation analysis of the protein coding genes detected indicates superior transcript expression correlation between low and high RNA inputs. This result demonstrates that D-Plex is an ideal solution for challenging samples such as FFPE preparations.</p>
</div>
</div>
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<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex UDI Module - Set C" target="_blank"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
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<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li>Ultra-low input capability, down to 10 pg for small RNAs and 100 pg for total RNA samples</li>
<li>High library complexity providing novel and diverse transcript detection</li>
<li>Versatile integartion into numerpus transcriptomic analysis workflow: ribosome profiling, CLIP-seq, RIP-seq and other</li>
<li>Improved quantification accuracy through the use of UMIs</li>
<li>Easy to use with minimal hands-on time: one day, one tube protocol</li>
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<p></p>
<center><em><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/MA-D-Plex.pdf" target="_blank" title="D-Plex Small RNA library preparation user manual"><strong>Download the manual</strong></a></em></center>
<p>D-Plex Small RNA-seq Library Prep Kit is a tool designed for the study of the <strong>small non-coding transcriptome</strong>. The kit is using the<a href="https://www.diagenode.com/en/pages/dplex" target="_blank"> D-Plex technology</a> to generate small RNA libraries for Illumina sequencing directly from total RNAs or enriched small RNAs.</p>
<p>The D-Plex technology utilizes two innovative <strong>ligation-free mechanisms</strong> - <strong>poly(A) tailing</strong> and <strong>template switching</strong> - to produce sequencing libraries from ultra-low input amounts, down to 10 pg for small RNAs and 100 pg for total RNAs. Combined with <strong>unique molecular identifiers</strong> (UMI), this complete solution ensures a <strong>realistic</strong> and <strong>accurate representation of diverse small RNA species</strong> (such as microRNAs, piRNAs, tRNAs, and siRNAs) with minimized quantitative bias.</p>
<p>D-Plex Small RNA-seq Kit offers a time saving protocol that can be completed within <strong>5 hours</strong> and requires minimal hands-on time. <span>The library preparation takes place in a </span><strong>single tube</strong><span>, increasing the efficiency tremendously. </span></p>
<p>D-Plex Small RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Indexes were designed and validated to fit the D-Plex technology and are available separately:</p>
<p><strong>For D-Plex UDI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><strong>For D-Plex SI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-A" title="D-Plex SI Module - Set A" target="_blank">C05030010 - D-Plex Single Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-B" title="D-Plex SI Module - Set B" target="_blank">C05030011 - D-Plex Single Indexes for Illumina - Set B</a></li>
</ul>
<p><strong>D-Plex is also available for MGI sequencing, check <a href="https://www.diagenode.com/en/p/d-plex-small-RNA-dnbseq-kit-x24" target="_blank">here</a>!</strong></p>
<p></p>
<p></p>
<p></p>
<center><strong>The D-Plex smRNA-seq is a versatile tool for a plethora of transciptomic analysis</strong></center>
<p></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/versatile-circle.png" /></center>
<p></p>
<p>With low-input capability, accuracy and ability to incorporate different transcript classes independent of chemical moieties, the strong D-Plex library-preparation method can be included in various analysis workflows (i.e., ribosome profiling, genome-wide mapping of protein: RNA binding sites).</p>
<p></p>
<p></p>',
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<div class="small-12 columns">
<p><strong>High library diversity for ultra-low inputs</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-diversity-fig1.png" /></center></div>
<div class="small-12 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig2-captures.png" /></center></div>
</div>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>D-Plex smRNA-seq captures the smRNA complexity of the sample in a sensitive manner. A large number of features is detected for each biotype at a CPM ≥5, as shown above for different species.</p>
</div>
</div>
<p></p>
<p><strong>Accurate representation of the smRNA content of a sample</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-ratplasma.png" /></center></div>
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-miRNAdistribution.png" /></center></div>
</div>
<div class="row">
<div class="small-6 columns">
<p><strong>Accurate identification of PCR duplicates in low-input samples using unique molecular identifiers (UMIs).</strong><br />The UMI read deduplication that is embedded in the D-Plex construct is used to avoid over-estimation of transcripts by identifying clones generated during the PCR amplification of the library. Despite limiting starting RNA input, the UMI correction removed technical copies of transcripts, maintaining the initial sample abundance.</p>
</div>
<div class="small-6 columns">
<p>D-Plex (template switching) technology enables recovery of the initial miRNA distribution of the miRXplore Universal Reference (Miltenyi Biotech Inc., Cat. No. 130-093-521). In contrast, ligation-based kits display a strong distortion of the miRNA equimolar distribution initially present in the pool, indicating sample incorporation bias.</p>
</div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-foldchange.jpg" /></center></div>
<div class="small-6 columns"><strong>D-Plex smRNA-seq in association with the MGcount analysis correlates strongly (R² > 0.8) with the RT-qPCR analysis, which is considered the gold-standard for accuracy. </strong><br />The figures shows a fold-change accuracy of a panel of 20 different small-RNA (figure taken from MGcount publication - Hita et al., BMC bioinformatics, 23-39(2022).</div>
</div>
<p></p>
<p></p>
<p><strong>Maximize information from the regulatory transcriptome using D-Plex small RNA-seq kit and MGcount*</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-multialignments.png" /></center></div>
<div class="small-6 columns">By analyzing D-Plex dataset with MGcount, more reads are kept during the analysis, which ultimately preserves biological complexity linked to ncRNA expression without decreasing the accuracy of detection. <br /><span style="line-height: 1em;"><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></span></div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>The D-Plex technology uses two innovative ligation-free mechanisms, <strong>poly(A) tailing and template switching</strong>, to produce sequencing libraries from ultra-low input amounts.</p>
</div>
</div>
<div class="row" style="background-color: #f5f5f5;">
<div class="small-12 columns"><br />
<p><strong>WORKFLOW</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/FL-Image-Workflow.jpg" width="50%" /></center></div>
<div class="small-12 columns">
<p style="text-align: justify; padding: 15px;"><br />RNA molecules are polyadenylated at the 3’-end and primed with an oligo(dT) primer containing part of the ILMN 3’-adapter sequence. The addition of a template-switching oligonucleotide during cDNA synthesis enables fusion of part of the ILMN 5’-adapter during reverse transcription. After PCR, the library comprises double stranded DNA constructs that are ready for sequencing.</p>
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<p><br /><br /></p>
<p></p>
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'info2' => '<p>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
<p>Need help for your bioinformatics analysis of miRNA?</p>
<p></p>
<div class="small-12 medium-3 large-3 columns"><center><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"><img src="https://www.diagenode.com/img/product/kits/dplex/miRMaster_logo.png" /></a></center>
<p> </p>
<p> </p>
</div>
<div class="small-12 medium-9 large-9 columns">
<p>Diagenode has collaborated with <strong>Andreas Keller</strong> and <strong>Tobias Fehlmann</strong> to develop a new bioinformatics pipeline for <span>the <strong>prediction of novel miRNAs</strong>.</span><br /><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"></a></p>
<ul>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster2">miRMaster 2</a></li>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank">miRMaster</a></li>
</ul>
</div>
<p> </p>
<p> </p>',
'label3' => 'Data analysis - Counting using MGcount',
'info3' => '<p>The counting or expression level calculation is the last step of the processing to generate an expression level matrix. We recommend using MGcount*, a counting tool developed at Diagenode with a suitable annotations file that include the small RNA transcripts that are object of study. MGcount, built on top of featureCounts, employs a flexible quantification approach to deal with datasets containing multiple RNA biotypes such as D-plex libraries. Non-coding RNAs varying in length, biogenesis, and function, may overlap in a genomic region, and are sometimes present in the genome with a high copy number. Consequently, reads may align equally well to more than one position in the reference genome or/and align to a position where more than one annotated transcript is located. MGcount employs two strategies to quantify these reads. Firstly, it assigns reads in rounds prioritizing small-RNAs over long-RNAs ensuring the unbiased read attribution to intergenic and intragenic small RNAs while preventing host-genes transcription over-estimation. Secondly, MGcount collapses loci where reads consistently multi-map into communities of loci with a graph-based approach. The resultant communities are groups of biologically related loci with nearly identical sequence. Subsequently, quantification of reads is performed at the loci-community level, reducing multi-alignments ambiguity at individual locus. Ultimately, this strategy maximizes the transcriptome information analysed and improves the interrogation of non-coding RNAs. MGcount software repository provides annotation files for A. thaliana, H. sapiens, M. musculus and C. elegans. The required input files for MGcount are a .txt file listing the paths to the alignment input files (.bam format) and the annotations file (.gtf format). The output directory path has to be provided as an input as well.</p>
<p><strong>Example command:</strong></p>
<p style="background-color: #f0f0f0;">MGcount -T 2 --gtf Homo_sapiens.GRCh38.gtf --outdir outputs --bam_infiles input_bamfilenames.txt</p>
<p>MGcount provides the choice to enable/disable the quantification of all RNA biotypes included in the annotation file in the form of "communities" as optional parameters for small-RNA (--ml_flag_small) and long-RNA (--ml_flag_long). Both are enabled by default. The main output of MGcount is the count_matrix.csv file containing an expression matrix that can be imported to R or any other software for downstream analyses.<br />A full user guide for MGCount is available here: <a href="https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html">https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html</a>.</p>
<p>Other standard counting tools such as featureCounts or HTSeq-counts can also be used alternatively. Given the high complexity of D-Plex libraries, we recommend having a clear understanding of the scientific question and the goal of the project before proceeding to the choice of the counting method as this will strongly impact downstream analyses. Diagenode provides bioinformatics support as a service (please, contact us if you need help with data analysis).</p>
<p>Notice that D-Plex produces forward-stranded data. Stranded libraries have the benefit that reads map to the genome strand where they were originated from. Therefore, when estimating transcript expression, reads aligned to the forward strand should be assigned only to transcript features in the forward strand whereas reads aligned to the reverse strand should be assigned only to transcript features in the reverse strand. For this, make sure you select “stranded mode” in any tool of choice. Stranded mode is selected by default in MGcount.</p>
<p><em><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></em></p>',
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'slug' => 'D-Plex-Small-RNA-seq-Library-Prep-x24',
'meta_title' => 'D-Plex Small RNA-seq Library Prep Kit | Diagenode ',
'meta_keywords' => 'RNA kit, RNA-seq kit, small RNA kit, small RNA-seq kit, low input RNA-seq, UMI RNA-seq, RNA-seq library preparation, D-Plex, higher RNA diversity',
'meta_description' => 'Small RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (100 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with plasma samples - User-friendly and fast protocol',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set A includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li>C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex 24 Single Indexes for Illumina - Set #B">C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex 24 Single Indexes for Illumina - Set #C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex 24 Single Indexes for Illumina - Set #D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<li>Compatible with<span> </span><a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24">D-Plex Small RNA-seq kit for Illumina</a> </li>
<li>Multiplexing up to <strong>96 samples</strong> when combining Set A, B, C and D</li>
<li>Allows for identification of index hopping</li>
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'info2' => '<p><span>D-Plex RNA-seq UDI library constructs bear the TruSeq (Illumina) HT adapters. In case of a multiplexing scenario, it is therefore recommended to submit the D-Plex libraries as TruSeq HT libraries to your sequencing provider. </span><span>Further details are provided in the D-Plex Unique Dual Indexes Module<span> </span><a href="https://www.diagenode.com/files/products/kits/dplex-unique-dual-indexes-manual.pdf" target="_blank">manual</a>.</span></p>',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A">C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li>C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<li>Multiplexing up to <strong>96 samples</strong> when combining Set A, B, C and D</li>
<li>Allows for identification of index hopping</li>
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<div class="small-12 medium-12 large-12 columns">
<h2 style="font-size: 22px;">DNA断片化、ライブラリー調製、自動化：NGSのワンストップショップ</h2>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-12 medium-12 large-12 columns">
<h4>1. 断片化装置を選択してください：150 bp〜75 kbの範囲でDNAを断片化します。</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-pico-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/bioruptor_pico.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/megaruptor2-1-unit"><img src="https://www.diagenode.com/img/product/shearing_technologies/B06010001_megaruptor2.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-one-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/br-one-profil.png" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns">5μlまで断片化：150 bp〜2 kb<br />NGS DNAライブラリー調製およびFFPE核酸抽出に最適で、</td>
<td class="small-4 medium-4 large-4 columns">2 kb〜75 kbの範囲をできます。<br />メイトペアライブラリー調製および長いフラグメントDNAシーケンシングに最適で、この軽量デスクトップデバイスで</td>
<td class="small-4 medium-4 large-4 columns">20または50μlの断片化が可能です。</td>
</tr>
</tbody>
</table>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-8 medium-8 large-8 columns">
<h4>2. 最適化されたライブラリー調整キットを選択してください。</h4>
</th>
<th class="small-4 medium-4 large-4 columns">
<h4>3. ライブラリー前処理自動化を選択して、比類のないデータ再現性を実感</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><img src="https://www.diagenode.com/img/product/kits/microPlex_library_preparation.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/ideal-library-preparation-kit-x24-incl-index-primer-set-1-24-rxns"><img src="https://www.diagenode.com/img/product/kits/box_kit.jpg" style="display: block; margin-left: auto; margin-right: auto;" height="173" width="250" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/sx-8g-ip-star-compact-automated-system-1-unit"><img src="https://www.diagenode.com/img/product/automation/B03000002%20_ipstar_compact.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">50pgの低入力：MicroPlex Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">5ng以上：iDeal Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">Achieve great NGS data easily</td>
</tr>
</tbody>
</table>
</div>
</div>
<blockquote>
<div class="row">
<div class="small-12 medium-12 large-12 columns"><span class="label" style="margin-bottom: 16px; margin-left: -22px; font-size: 15px;">DiagenodeがNGS研究にぴったりなプロバイダーである理由</span>
<p>Diagenodeは15年以上もエピジェネティクス研究に専念、専門としています。 ChIP研究クロマチン用のユニークな断片化システムの開発から始まり、 専門知識を活かし、5μlのせん断体積まで可能で、NGS DNAライブラリーの調製に最適な最先端DNA断片化装置の開発にたどり着きました。 我々は以来、ChIP-seq、Methyl-seq、NGSライブラリー調製用キットを研究開発し、業界をリードする免疫沈降研究と同様に、ライブラリー調製を自動化および完結させる独自の自動化システムを開発にも成功しました。</p>
<ul>
<li>信頼されるせん断装置</li>
<li>様々なインプットからのライブラリ作成キット</li>
<li>独自の自動化デバイス</li>
</ul>
</div>
</div>
</blockquote>
<div class="row">
<div class="small-12 columns">
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#panel1a">次世代シーケンシングへの理解とその専門知識</a>
<div id="panel1a" class="content">
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<p><strong>次世代シーケンシング (NGS)</strong> ）は、著しいスケールとハイスループットでシーケンシングを行い、1日に数十億もの塩基生成を可能にします。 NGSのハイスループットは迅速でありながら正確で、再現性のあるデータセットを実現し、さらにシーケンシング費用を削減します。 NGSは、ゲノムシーケンシング、ゲノム再シーケンシング、デノボシーケンシング、トランスクリプトームシーケンシング、その他にDNA-タンパク質相互作用の検出やエピゲノムなどを示します。 指数関数的に増加するシーケンシングデータの需要は、計算分析の障害や解釈、データストレージなどの課題を解決します。</p>
<p>アプリケーションおよび出発物質に応じて、数百万から数十億の鋳型DNA分子を大規模に並行してシーケンシングすることが可能です。その為に、異なる化学物質を使用するいくつかの市販のNGSプラットフォームを利用することができます。 NGSプラットフォームの種類によっては、事前準備とライブラリー作成が必要です。</p>
<p>NGSにとっても、特にデータ処理と分析に関した大きな課題はあります。第3世代技術はゲノミクス研究にさらに革命を起こすであろうと大きく期待されています。</p>
</div>
</div>
<div class="row">
<div class="small-6 medium-6 large-6 columns">
<p><strong>NGS アプリケーション</strong></p>
<ul>
<li>全ゲノム配列決定</li>
<li>デノボシーケンシング</li>
<li>標的配列</li>
<li>Exomeシーケンシング</li>
<li>トランスクリプトーム配列決定</li>
<li>ゲノム配列決定</li>
<li>ミトコンドリア配列決定</li>
<li>DNA-タンパク質相互作用（ChIP-seq</li>
<li>バリアント検出</li>
<li>ゲノム仕上げ</li>
</ul>
</div>
<div class="small-6 medium-6 large-6 columns">
<p><strong>研究分野におけるNGS:</strong></p>
<ul>
<li>腫瘍学</li>
<li>リプロダクティブ・ヘルス</li>
<li>法医学ゲノミクス</li>
<li>アグリゲノミックス</li>
<li>複雑な病気</li>
<li>微生物ゲノミクス</li>
<li>食品・環境ゲノミクス</li>
<li>創薬ゲノミクス - パーソナライズド・メディカル</li>
</ul>
</div>
<div class="small-12 medium-12 large-12 columns">
<p><strong>NGSの用語</strong></p>
<dl>
<dt>リード（読み取り）</dt>
<dd>この装置から得られた連続した単一のストレッチ</dd>
<dt>断片リード</dt>
<dd>フラグメントライブラリからの読み込み。 シーケンシングプラットフォームに応じて、読み取りは通常約100〜300bp。</dd>
<dt>断片ペアエンドリード</dt>
<dd>断片ライブラリーからDNA断片の各末端2つの読み取り。</dd>
<dt>メイトペアリード</dt>
<dd>大きなDNA断片（通常は予め定義されたサイズ範囲）の各末端から2つの読み取り。</dd>
<dt>カバレッジ（例）</dt>
<dd>30×適用範囲とは、参照ゲノム中の各塩基対が平均30回の読み取りを示す。</dd>
</dl>
</div>
</div>
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2>NGSプラットフォーム</h2>
<h3><a href="http://www.illumina.com" target="_blank">イルミナ</a></h3>
<p>イルミナは、クローン的に増幅された鋳型DNA（クラスター）上に位置する、蛍光標識された可逆的鎖ターミネーターヌクレオチドを用いた配列別合成技術を使用。 DNAクラスターは、ガラスフローセルの表面上に固定化され、 ワークフローは、4つのヌクレオチド（それぞれ異なる蛍光色素で標識された）の組み込み、4色イメージング、色素や末端基の切断、取り込み、イメージングなどを繰り返します。フローセルは大規模な並列配列決定を受ける。 この方法により、単一蛍光標識されたヌクレオチドの制御添加によるモノヌクレオチドのエラーを回避する可能性があります。 読み取りの長さは、通常約100〜150 bpです。</p>
<h3><a href="http://www.lifetechnologies.com" target="_blank">イオン トレント</a></h3>
<p>イオントレントは、半導体技術チップを用いて、合成中にヌクレオチドを取り込む際に放出されたプロトンを検出します。 これは、イオン球粒子と呼ばれるビーズの表面にエマルションPCR（emPCR）を使用し、リンクされた特定のアダプターを用いてDNA断片を増幅します。 各ビーズは1種類のDNA断片で覆われていて、異なるDNA断片を有するビーズは次いで、チップの陽子感知ウェル内に配置されます。 チップには一度に4つのヌクレオチドのうちの1つが浸水し、このプロセスは異なるヌクレオチドで15秒ごとに繰り返されます。 配列決定の間に4つの塩基の各々が1つずつ導入されます、組み込みの場合はプロトンが放出され、電圧信号が取り込みに比例して検出されます。.</p>
<h3><a href="http://www.pacificbiosciences.com" target="_blank">パシフィック バイオサイエンス</a></h3>
<p>パシフィックバイオサイエンスでは、20kbを超える塩基対の読み取りも、単一分子リアルタイム（SMRT）シーケンシングによる構造および細胞タイプの変化を観察することができます。 このプラットフォームでは、超長鎖二本鎖DNA（dsDNA）断片が、Megaruptor（登録商標）のようなDiagenode装置を用いたランダムシアリングまたは目的の標的領域の増幅によって生成されます。 SMRTbellライブラリーは、ユニバーサルヘアピンアダプターをDNA断片の各末端に連結することによって生成します。 サイズ選択条件による洗浄ステップの後、配列決定プライマーをSMRTbellテンプレートにアニーリングし、鋳型DNAに結合したDNAポリメラーゼを含む配列決定を、蛍光標識ヌクレオチドの存在下で開始。 各塩基が取り込まれると、異なる蛍光のパルスをリアルタイムで検出します。</p>
<h3><a href="https://nanoporetech.com" target="_blank">オックスフォード ナノポア</a></h3>
<p>Oxford Nanoporeは、単一のDNA分子配列決定に基づく技術を開発します。その技術により生物学的分子、すなわちDNAが一群の電気抵抗性高分子膜として位置するナノスケールの孔（ナノ細孔）またはその近くを通過し、イオン電流が変化します。 この変化に関する情報は、例えば4つのヌクレオチド（AまたはG r CまたはT）ならびに修飾されたヌクレオチドすべてを区別することによって分子情報に訳されます。 シーケンシングミニオンデバイスのフローセルは、数百個のナノポアチャネルのセンサアレイを含みます。 DNAサンプルは、Diagenode社のMegaruptor（登録商標）を用いてランダムシアリングによって生成され得る超長鎖DNAフラグメントが必要です。</p>
<h3><a href="http://www.lifetechnologies.com/be/en/home/life-science/sequencing/next-generation-sequencing/solid-next-generation-sequencing.html" target="_blank">SOLiD</a></h3>
<p>SOLiDは、ユニークな化学作用により、何千という個々のDNA分子の同時配列決定を可能にします。 それは、アダプター対ライブラリーのフラグメントが適切で、せん断されたゲノムDNAへのアダプターのライゲーションによるライブラリー作製から始まります。 次のステップでは、エマルジョンPCR（emPCR）を実施して、ビーズの表面上の個々の鋳型DNA分子をクローン的に増幅。 emPCRでは、個々の鋳型DNAをPCR試薬と混合し、水中油型エマルジョン内の疎水性シェルで囲まれた水性液滴内のプライマーコートビーズを、配列決定のためにロードするスライドガラスの表面にランダムに付着。 この技術は、シークエンシングプライマーへのライゲーションで競合する4つの蛍光標識されたジ塩基プローブのセットを使用します。</p>
<h3><a href="http://454.com/products/technology.asp" target="_blank">454</a></h3>
<p>454は、大規模並列パイロシーケンシングを利用しています。 始めに全ゲノムDNAまたは標的遺伝子断片の300〜800bp断片のライブラリー調製します。 次に、DNAフラグメントへのアダプターの付着および単一のDNA鎖の分離。 その後アダプターに連結されたDNAフラグメントをエマルジョンベースのクローン増幅（emPCR）で処理し、DNAライブラリーフラグメントをミクロンサイズのビーズ上に配置します。 各DNA結合ビーズを光ファイバーチップ上のウェルに入れ、器具に挿入します。 4つのDNAヌクレオチドは、配列決定操作中に固定された順序で連続して加えられ、並行して配列決定されます。</p>
</div>
</div>
</div>
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<p><span style="font-weight: 400;">Most of the major next-generation sequencing platforms require ligation of specific adaptor oligos to </span><a href="../applications/dna-rna-shearing"><span style="font-weight: 400;">fragmented DNA or RNA</span></a><span style="font-weight: 400;"> prior to sequencing</span></p>
<p><span style="font-weight: 400;">After input DNA has been fragmented, it is end-repaired and blunt-ended</span><span style="font-weight: 400;">. The next step is a A-tailing in which dAMP is added to the 3´ end of the blunt phosphorylated DNA fragments to prevent concatemerization and to allow the ligation of adaptors with complementary dT overhangs. In addition, barcoded adapters can be incorporated to facilitate multiplexing prior to or during amplification.</span></p>
<center><img src="https://www.diagenode.com/img/categories/library-prep/flux.png" /></center>
<p><span style="font-weight: 400;">Diagenode offers a comprehensive product portfolio for library preparation:<br /></span></p>
<strong><a href="https://www.diagenode.com/en/categories/Library-preparation-for-RNA-seq">D-Plex RNA-seq Library Preparation Kits</a></strong><br />
<p><span style="font-weight: 400;">Diagenode’s new RNA-sequencing solutions utilize the innovative c</span><span style="font-weight: 400;">apture and a</span><span style="font-weight: 400;">mplification by t</span><span style="font-weight: 400;">ailing and s</span><span style="font-weight: 400;">witching”</span><span style="font-weight: 400;">, a ligation-free method to produce DNA libraries for next generation sequencing from low input amounts of RNA. </span><span style="font-weight: 400;"></span><a href="../categories/Library-preparation-for-RNA-seq">Learn more</a></p>
<strong><a href="../categories/library-preparation-for-ChIP-seq">ChIP-seq and DNA sequencing library preparation solutions</a></strong><br />
<p><span style="font-weight: 400;">Our kits have been optimized for DNA library preparation used for next generation sequencing for a wide range of inputs. Using a simple three-step protocols, our</span><a href="http://www.diagenode.com/p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><span style="font-weight: 400;"> </span></a><span style="font-weight: 400;">kits are an optimal choice for library preparation from DNA inputs down to 50 pg. </span><a href="../categories/library-preparation-for-ChIP-seq">Learn more</a></p>
<a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span><strong>Bioruptor Pico - short fragments</strong></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">Our well-cited Bioruptor Pico is the shearing device of choice for chromatin and DNA fragmentation. Obtain uniform and tight fragment distributions between 150bp -2kb. </span><a href="../p/bioruptor-pico-sonication-device">Learn more</a></p>
<strong><a href="../p/megaruptor2-1-unit"><span href="../p/bioruptor-pico-sonication-device">Megaruptor</span>® - long fragments</a></strong><a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">The Megaruptor is designed to shear DNA from 3kb-75kb for long-read sequencing. <a href="../p/megaruptor2-1-unit">Learn more</a></span></p>
<span href="../p/bioruptor-pico-sonication-device"></span><span style="font-weight: 400;"></span></div>
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'description' => '<p><span>Nutrient scarcity is a frequent adverse condition that organisms face during their development. This condition may lead to long-lasting effects on the metabolism and behaviour of adults due to developmental epigenetic modifications. Here, we show that reducing nutrient availability during larval development affects adult spontaneous activity and sleep behaviour, together with changes in gene expression and epigenetic marks in the mushroom bodies (MBs). We found that open chromatin regions map to 100 of 241 transcriptionally upregulated genes in the adult MBs, these new opening zones are preferentially located in regulatory zones such as promoter-TSS and introns. Importantly, opened chromatin at the Dopamine 1-like receptor 2 regulatory zones correlate with increased expression. In consequence, adult administration of a dopamine antagonist reverses increased spontaneous activity and diminished sleep time observed in response to early-life nutrient restriction. In comparison, reducing </span><i>dop1R2</i><span><span> </span>expression in MBs also ameliorates these effects, albeit to a lesser degree. These results lead to the conclusion that increased dopamine signalling in the MBs of flies reared in a poor nutritional environment underlies the behavioural changes observed due to this condition during development.</span></p>',
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'doi' => '10.1098/rsob.230049',
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'authors' => 'Garza A. et al.',
'description' => '<p><span>Cobalt and Nickel show increasing pollution in our environment. We analyzed their combined toxicity and uptake mechanisms in the early food chain by investigation of bacteria and the ciliate Paramecium as a primary consumer. Exposing Paramecium to metals, we can dissect between phagocytotic uptake and phagocytosis independent uptake. Measurement of cellular content in bacteria after exposure shows a bias for Nickel when both are co-exposed and even increased levels in comparison to single exposure. Imitating the initial food chain, we fed these bacteria to paramecia measuring only phagocytotic uptake. The cellular content shows a similar ratio of Nickel and Cobalt as in food bacteria suggesting that bacteria can selectively pre-accumulate metals for introduction into the food-chain. Analyzing the transcriptomic response of paramecia to sublethal doses of Nickel and Cobalt, Gene ontogogy (GO)-analysis indicates common deregulated pathways, e.g. the ammonium-transmembrane transport and ubiquitine-associated protein degradation. Redox related genes show a massive de-regulation of gene expression indicating cellular adaptation to increased ROS stress. As this suggests that both metals can target also the same cellular pathways, this is in agreement with increased toxicity of both metals in joint applications. Our data reveals complex routes of individual metals to enter the food chain.</span></p>',
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'name' => 'Manganese-Induced Toxicity in : What Can We Learn from theTranscriptome?',
'authors' => 'Nicolai Merle M et al.',
'description' => '<p>Manganese (Mn) is an essential ubiquitous transition metal and, when occupationally or environmentally overexposed, a well-known risk factor for several neurological pathologies. However, the molecular mechanisms underlying Mn-induced neurotoxicity are largely unknown. In this study, addressing RNA-Seq analysis, bioavailability and survival assays, key pathways of transcriptional responses to Mn overexposure were investigated in the model organism (), providing insights into the Mn-induced cellular stress and damage response. Comparative transcriptome analyses identified a large number of differentially expressed genes (DEGs) in nematodes exposed to MnCl, and functional annotation suggested oxidative nucleotide damage, unfolded protein response and innate immunity as major damage response pathways. Additionally, a time-dependent increase in the transcriptional response after MnCl exposure was identified by means of increased numbers of DEGs, indicating a time-dependent response and activation of the stress responses in Mn neurotoxicity. The data provided here represent a powerful transcriptomic resource in the field of Mn toxicity, and therefore, this study provides a useful basis for further planning of targeted mechanistic studies of Mn-induced neurotoxicity that are urgently needed in the face of increasing industrially caused environmental pollution with Mn.</p>',
'date' => '2022-09-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/36142660',
'doi' => '10.3390/ijms231810748',
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'name' => 'A Chromosomal Duplication Encompassing Interleukin-33 Causes aNovel Hyper IgE Phenotype Characterized by EosinophilicEsophagitis and Generalized Autoimmunity.',
'authors' => 'Marwaha A.K. et al.',
'description' => '<p>Interleukin (IL)-33 is a member of the IL-1 cytokine family and is secreted by fibroblasts, endothelial cells, and epithelial cells in tissues (lung, skin, and gastrointestinal) that are exposed to the environment. 1 IL-33 is stored in the nucleus and environmental allergens trigger RIPK1-caspase 8 ripoptosome activation in epithelial cells to enable IL-33 maturation and release. 2 IL-33 signals via its receptor ST2, which is found on multiple immune cells, such as eosinophils, mast cells, T cells, macrophages, basophils, and type 2 innate lymphoid cells, which all promote type-2 innate immune reactions. 3 Therefore, IL-33 plays a central role in promoting allergic inflammatory responses in diseases such as eosinophilic esophagitis (EoE), allergy to food/inhalants, asthma, and atopic dermatitis. Genome-wide association data of these conditions consistently report polymorphisms in the IL-33 gene as a risk factor. 4 Thus, IL-33 has been proposed as a therapeutic target for these conditions. In animal models, IL-33 has been mechanistically linked with allergic, autoimmune, rheumatologic, and inflammatory bowel diseases. 5 However, monogenic human diseases caused by either gain-of-function or loss-of-function variants in the IL-33 gene have not been reported previously.</p>',
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'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/35489436',
'doi' => '10.1053/j.gastro.2022.04.026',
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'name' => 'Functional microRNA targetome undergoes degeneration-induced shift inthe retina.',
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'description' => '<p>BACKGROUND: MicroRNA (miRNA) play a significant role in the pathogenesis of complex neurodegenerative diseases including age-related macular degeneration (AMD), acting as post-transcriptional gene suppressors through their association with argonaute 2 (AGO2) - a key member of the RNA Induced Silencing Complex (RISC). Identifying the retinal miRNA/mRNA interactions in health and disease will provide important insight into the key pathways miRNA regulate in disease pathogenesis and may lead to potential therapeutic targets to mediate retinal degeneration. METHODS: To identify the active miRnome targetome interactions in the healthy and degenerating retina, AGO2 HITS-CLIP was performed using a rodent model of photoreceptor degeneration. Analysis of publicly available single-cell RNA sequencing (scRNAseq) data was performed to identify the cellular location of AGO2 and key members of the microRNA targetome in the retina. AGO2 findings were verified by in situ hybridization (RNA) and immunohistochemistry (protein). RESULTS: Analysis revealed a similar miRnome between healthy and damaged retinas, however, a shift in the active targetome was observed with an enrichment of miRNA involvement in inflammatory pathways. This shift was further demonstrated by a change in the seed binding regions of miR-124-3p, the most abundant retinal AGO2-bound miRNA, and has known roles in regulating retinal inflammation. Additionally, photoreceptor cluster miR-183/96/182 were all among the most highly abundant miRNA bound to AGO2. Following damage, AGO2 expression was localized to the inner retinal layers and more in the OLM than in healthy retinas, indicating a locational miRNA response to retinal damage. CONCLUSIONS: This study provides important insight into the alteration of miRNA regulatory activity that occurs as a response to retinal degeneration and explores the miRNA-mRNA targetome as a consequence of retinal degenerations. Further characterisation of these miRNA/mRNA interactions in the context of the degenerating retina may provide an important insight into the active role these miRNA may play in diseases such as AMD.</p>',
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'description' => '<div class="row"><div class="small-12 medium-4 large-4 columns"><div class="panel"><h3><em>lncRNAs in cancer</em></h3><p class="text-left">Gastrointestinal cancer: Gastrointestinal cancers occur as a result of dysregulated signaling pathways and cellular processes, such as the cell cycle or apoptosis. LncRNAs can regulate proliferation of gastrointestinal cancer through interacting with RNA targets, localization to chromatin, or binding to proteins. Read more about Long non-coding RNAs: crucial regulators of gastrointestinal cancer cell proliferation</p><h3><em>microRNAs in cancer</em></h3><p class="text-left">miR-155 has been found overexpressed in many cancer types including hematopoietic cancers, breast, lung and colon cancer<br /><br /> <a href="https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf">https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf</a></p></div></div><div class="small-12 medium-8 large-8 columns"><center><img src="https://www.diagenode.com/img/cancer/long-non-coding-rna.jpg" width="550" height="345" /></center><p></p><p>Various non-coding RNAs (ncRNAs) have been found to function as key regulators for transcription, chromatin remodelling, and post-transcriptional modification, thus making them relevant in oncogenesis, both as tumor suppressors and as drivers. For example, a number of microRNAs (miRNAs), one of the more well-studied types of ncRNAs, have been identified as potential cancer biomarkers and clinical targets. Other ncRNAs, such as long noncoding RNAs are involved in cancer regulation and proliferation. Understanding the role of ncRNAs will be critical for cancer research and therapeutic development.</p></div></div>',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
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<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li><span style="font-weight: 400;">An innovative technology with template switching and UMIs</span></li>
<li><span style="font-weight: 400;">Ultra-low input capability, down to 50 pg for total RNAs</span></li>
<li><span style="font-weight: 400;">Capture the widest possible diversity of RNAs for rich content</span></li>
<li><span>Get high sensitivity data even from difficult samples, such as degraded, FFPE samples</span></li>
<li><span style="font-weight: 400;">Enjoy a fast, easy, single tube protocol</span></li>
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<p></p>
<p></p>
<h4></h4>
<center><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/dplex-total-manual.pdf" target="_blank" title="dplex total rnaseq user manual"><strong>Download the manual</strong></a></center>
<p></p>
<p>D-Plex Total RNA-seq Library Preparation Kit is a tool designed for the study of the whole coding and non-coding transcriptome. <span>The kit is using the</span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank"><span> </span>D-Plex technology</a><span><span> </span>to generate directional libraries for Illumina sequencing directly from total RNAs, mRNAs that has already been enriched by poly(A) selection, or RNAs that has already been depleted of rRNAs.</span></p>
<p><span>The D-Plex technology utilizes two innovative ligation-free mechanisms - poly(A) tailing and template switching - to produce sequencing libraries from ultra-low input amounts, down to 50 pg for total RNAs, mRNAs or rRNA-depleted RNAs. Combined with unique molecular identifiers (UMI), this complete solution delivers a high-sensitivity detection method for comprehensive analysis of the transcriptome. It accurately measures gene and transcript abundance and captures the widest possible diversity of RNAs, including known and novel features in coding and non-coding RNAs.</span></p>
<p><span></span><span>D-Plex Total RNA-seq Kit offers a time saving protocol that can be completed within 5 hours and requires minimal hands-on time. <span>The library preparation takes place in a </span><span>single tube</span><span>, increasing the efficiency tremendously. This new solution has been extensively validated for both intact and highly degraded RNA samples, including that derived from FFPE preparations.</span></span></p>
<p><span><span>D-Plex Total RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Unique Dual Indexes were designed and validated to fit the D-Plex technology and are available separately:</span></span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C </a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<div class="extra-spaced" align="center"></div>
<div class="row">
<div class="carrousel" style="background-position: center;">
<div class="slick">
<div>
<h3>Greater sensitivity to detect novel transcripts</h3>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/CPM_totalrna.png" alt="total RNA kit" /></center></div>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq kit enables great transcript detection, even when starting from very low RNA inputs or working with challenging FFPE samples. Increased number of transcripts detected at 1x coverage is an indicator of greater sensitivity.</p>
</div>
</div>
<div>
<h3>Highest diversity to facilitate biomarker discovery</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_rdep_biotypes.png" alt="total RNA diversity" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq libraries capture efficiently all RNA biotypes present in the sample of interest, both coding and non-coding RNAs or small and long RNAs.</p>
</div>
</div>
<div>
<h3>Consistent expression profiling across a wide range of RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_and_umeg_rdep_violin_tpm_10.png" alt="total RNA" width="450px" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Gene expression profiling (including protein coding exons and introns) obtained with D-Plex Total RNA-seq kit is conserved for decreasing RNA inputs, keeping transcript diversity across the range of RNA inputs.</p>
</div>
</div>
<div>
<h3>Superior library complexity at low RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/robustness_corr_HUR_HuMEg_rrna.png" alt="total RNA" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Correlation analysis of the protein coding genes detected indicates superior transcript expression correlation between low and high RNA inputs. This result demonstrates that D-Plex is an ideal solution for challenging samples such as FFPE preparations.</p>
</div>
</div>
</div>
</div>
</div>',
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'label2' => 'Indexes',
'info2' => '<p><span>Specific D-Plex indexes </span><span>were designed and validated to fit the D-Plex technology for Illumina sequencing and </span><span>are not included in the kit. They can be bought separately according to your needs. Please choose the format that suits you best among the compatible references to:</span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex UDI Module - Set C" target="_blank"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<p></p>
<p>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</p>',
'label3' => 'Data analysis',
'info3' => '<p><span>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</span></p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="Small RNA seq Bioinformatics pipeline" width="925" height="196" /></p>',
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'slug' => 'D-Plex-Total-RNA-seq-Library-Prep-x24',
'meta_title' => 'D-Plex Total RNA-seq Library Prep Kit for Illumina | Diagenode ',
'meta_keywords' => 'RNA-seq kit, RNA-seq library preparation; low input RNA-seq, UMI RNA-seq, UDI RNA-seq, D-Plex, higher RNA sensitivity',
'meta_description' => 'RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (50 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with FFPE samples - User-friendly and fast protocol',
'modified' => '2023-11-16 09:09:25',
'created' => '2021-04-13 14:10:40',
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'name' => 'D-Plex Small RNA-seq Kit for Illumina',
'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li>Ultra-low input capability, down to 10 pg for small RNAs and 100 pg for total RNA samples</li>
<li>High library complexity providing novel and diverse transcript detection</li>
<li>Versatile integartion into numerpus transcriptomic analysis workflow: ribosome profiling, CLIP-seq, RIP-seq and other</li>
<li>Improved quantification accuracy through the use of UMIs</li>
<li>Easy to use with minimal hands-on time: one day, one tube protocol</li>
</ul>
</div>
<p></p>
<p></p>
<center><em><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/MA-D-Plex.pdf" target="_blank" title="D-Plex Small RNA library preparation user manual"><strong>Download the manual</strong></a></em></center>
<p>D-Plex Small RNA-seq Library Prep Kit is a tool designed for the study of the <strong>small non-coding transcriptome</strong>. The kit is using the<a href="https://www.diagenode.com/en/pages/dplex" target="_blank"> D-Plex technology</a> to generate small RNA libraries for Illumina sequencing directly from total RNAs or enriched small RNAs.</p>
<p>The D-Plex technology utilizes two innovative <strong>ligation-free mechanisms</strong> - <strong>poly(A) tailing</strong> and <strong>template switching</strong> - to produce sequencing libraries from ultra-low input amounts, down to 10 pg for small RNAs and 100 pg for total RNAs. Combined with <strong>unique molecular identifiers</strong> (UMI), this complete solution ensures a <strong>realistic</strong> and <strong>accurate representation of diverse small RNA species</strong> (such as microRNAs, piRNAs, tRNAs, and siRNAs) with minimized quantitative bias.</p>
<p>D-Plex Small RNA-seq Kit offers a time saving protocol that can be completed within <strong>5 hours</strong> and requires minimal hands-on time. <span>The library preparation takes place in a </span><strong>single tube</strong><span>, increasing the efficiency tremendously. </span></p>
<p>D-Plex Small RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Indexes were designed and validated to fit the D-Plex technology and are available separately:</p>
<p><strong>For D-Plex UDI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><strong>For D-Plex SI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-A" title="D-Plex SI Module - Set A" target="_blank">C05030010 - D-Plex Single Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-B" title="D-Plex SI Module - Set B" target="_blank">C05030011 - D-Plex Single Indexes for Illumina - Set B</a></li>
</ul>
<p><strong>D-Plex is also available for MGI sequencing, check <a href="https://www.diagenode.com/en/p/d-plex-small-RNA-dnbseq-kit-x24" target="_blank">here</a>!</strong></p>
<p></p>
<p></p>
<p></p>
<center><strong>The D-Plex smRNA-seq is a versatile tool for a plethora of transciptomic analysis</strong></center>
<p></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/versatile-circle.png" /></center>
<p></p>
<p>With low-input capability, accuracy and ability to incorporate different transcript classes independent of chemical moieties, the strong D-Plex library-preparation method can be included in various analysis workflows (i.e., ribosome profiling, genome-wide mapping of protein: RNA binding sites).</p>
<p></p>
<p></p>',
'label1' => 'Characteristics and library prep. workflow ',
'info1' => '<div class="row">
<div class="small-12 columns">
<p><strong>High library diversity for ultra-low inputs</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-diversity-fig1.png" /></center></div>
<div class="small-12 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig2-captures.png" /></center></div>
</div>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>D-Plex smRNA-seq captures the smRNA complexity of the sample in a sensitive manner. A large number of features is detected for each biotype at a CPM ≥5, as shown above for different species.</p>
</div>
</div>
<p></p>
<p><strong>Accurate representation of the smRNA content of a sample</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-ratplasma.png" /></center></div>
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-miRNAdistribution.png" /></center></div>
</div>
<div class="row">
<div class="small-6 columns">
<p><strong>Accurate identification of PCR duplicates in low-input samples using unique molecular identifiers (UMIs).</strong><br />The UMI read deduplication that is embedded in the D-Plex construct is used to avoid over-estimation of transcripts by identifying clones generated during the PCR amplification of the library. Despite limiting starting RNA input, the UMI correction removed technical copies of transcripts, maintaining the initial sample abundance.</p>
</div>
<div class="small-6 columns">
<p>D-Plex (template switching) technology enables recovery of the initial miRNA distribution of the miRXplore Universal Reference (Miltenyi Biotech Inc., Cat. No. 130-093-521). In contrast, ligation-based kits display a strong distortion of the miRNA equimolar distribution initially present in the pool, indicating sample incorporation bias.</p>
</div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-foldchange.jpg" /></center></div>
<div class="small-6 columns"><strong>D-Plex smRNA-seq in association with the MGcount analysis correlates strongly (R² > 0.8) with the RT-qPCR analysis, which is considered the gold-standard for accuracy. </strong><br />The figures shows a fold-change accuracy of a panel of 20 different small-RNA (figure taken from MGcount publication - Hita et al., BMC bioinformatics, 23-39(2022).</div>
</div>
<p></p>
<p></p>
<p><strong>Maximize information from the regulatory transcriptome using D-Plex small RNA-seq kit and MGcount*</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-multialignments.png" /></center></div>
<div class="small-6 columns">By analyzing D-Plex dataset with MGcount, more reads are kept during the analysis, which ultimately preserves biological complexity linked to ncRNA expression without decreasing the accuracy of detection. <br /><span style="line-height: 1em;"><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></span></div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>The D-Plex technology uses two innovative ligation-free mechanisms, <strong>poly(A) tailing and template switching</strong>, to produce sequencing libraries from ultra-low input amounts.</p>
</div>
</div>
<div class="row" style="background-color: #f5f5f5;">
<div class="small-12 columns"><br />
<p><strong>WORKFLOW</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/FL-Image-Workflow.jpg" width="50%" /></center></div>
<div class="small-12 columns">
<p style="text-align: justify; padding: 15px;"><br />RNA molecules are polyadenylated at the 3’-end and primed with an oligo(dT) primer containing part of the ILMN 3’-adapter sequence. The addition of a template-switching oligonucleotide during cDNA synthesis enables fusion of part of the ILMN 5’-adapter during reverse transcription. After PCR, the library comprises double stranded DNA constructs that are ready for sequencing.</p>
</div>
</div>
<p><br /><br /></p>
<p></p>
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'label2' => 'Data analysis',
'info2' => '<p>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
<p>Need help for your bioinformatics analysis of miRNA?</p>
<p></p>
<div class="small-12 medium-3 large-3 columns"><center><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"><img src="https://www.diagenode.com/img/product/kits/dplex/miRMaster_logo.png" /></a></center>
<p> </p>
<p> </p>
</div>
<div class="small-12 medium-9 large-9 columns">
<p>Diagenode has collaborated with <strong>Andreas Keller</strong> and <strong>Tobias Fehlmann</strong> to develop a new bioinformatics pipeline for <span>the <strong>prediction of novel miRNAs</strong>.</span><br /><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"></a></p>
<ul>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster2">miRMaster 2</a></li>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank">miRMaster</a></li>
</ul>
</div>
<p> </p>
<p> </p>',
'label3' => 'Data analysis - Counting using MGcount',
'info3' => '<p>The counting or expression level calculation is the last step of the processing to generate an expression level matrix. We recommend using MGcount*, a counting tool developed at Diagenode with a suitable annotations file that include the small RNA transcripts that are object of study. MGcount, built on top of featureCounts, employs a flexible quantification approach to deal with datasets containing multiple RNA biotypes such as D-plex libraries. Non-coding RNAs varying in length, biogenesis, and function, may overlap in a genomic region, and are sometimes present in the genome with a high copy number. Consequently, reads may align equally well to more than one position in the reference genome or/and align to a position where more than one annotated transcript is located. MGcount employs two strategies to quantify these reads. Firstly, it assigns reads in rounds prioritizing small-RNAs over long-RNAs ensuring the unbiased read attribution to intergenic and intragenic small RNAs while preventing host-genes transcription over-estimation. Secondly, MGcount collapses loci where reads consistently multi-map into communities of loci with a graph-based approach. The resultant communities are groups of biologically related loci with nearly identical sequence. Subsequently, quantification of reads is performed at the loci-community level, reducing multi-alignments ambiguity at individual locus. Ultimately, this strategy maximizes the transcriptome information analysed and improves the interrogation of non-coding RNAs. MGcount software repository provides annotation files for A. thaliana, H. sapiens, M. musculus and C. elegans. The required input files for MGcount are a .txt file listing the paths to the alignment input files (.bam format) and the annotations file (.gtf format). The output directory path has to be provided as an input as well.</p>
<p><strong>Example command:</strong></p>
<p style="background-color: #f0f0f0;">MGcount -T 2 --gtf Homo_sapiens.GRCh38.gtf --outdir outputs --bam_infiles input_bamfilenames.txt</p>
<p>MGcount provides the choice to enable/disable the quantification of all RNA biotypes included in the annotation file in the form of "communities" as optional parameters for small-RNA (--ml_flag_small) and long-RNA (--ml_flag_long). Both are enabled by default. The main output of MGcount is the count_matrix.csv file containing an expression matrix that can be imported to R or any other software for downstream analyses.<br />A full user guide for MGCount is available here: <a href="https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html">https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html</a>.</p>
<p>Other standard counting tools such as featureCounts or HTSeq-counts can also be used alternatively. Given the high complexity of D-Plex libraries, we recommend having a clear understanding of the scientific question and the goal of the project before proceeding to the choice of the counting method as this will strongly impact downstream analyses. Diagenode provides bioinformatics support as a service (please, contact us if you need help with data analysis).</p>
<p>Notice that D-Plex produces forward-stranded data. Stranded libraries have the benefit that reads map to the genome strand where they were originated from. Therefore, when estimating transcript expression, reads aligned to the forward strand should be assigned only to transcript features in the forward strand whereas reads aligned to the reverse strand should be assigned only to transcript features in the reverse strand. For this, make sure you select “stranded mode” in any tool of choice. Stranded mode is selected by default in MGcount.</p>
<p><em><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></em></p>',
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'slug' => 'D-Plex-Small-RNA-seq-Library-Prep-x24',
'meta_title' => 'D-Plex Small RNA-seq Library Prep Kit | Diagenode ',
'meta_keywords' => 'RNA kit, RNA-seq kit, small RNA kit, small RNA-seq kit, low input RNA-seq, UMI RNA-seq, RNA-seq library preparation, D-Plex, higher RNA diversity',
'meta_description' => 'Small RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (100 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with plasma samples - User-friendly and fast protocol',
'modified' => '2024-04-12 11:41:21',
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'name' => 'D-Plex Unique Dual Indexes for Illumina - Set A',
'description' => '<p><a href="https://www.diagenode.com/files/products/kits/dplex-unique-dual-indexes-manual.pdf"><img src="https://www.diagenode.com/img/buttons/bt-manual.png" /></a></p>
<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set A includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li>C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex 24 Single Indexes for Illumina - Set #B">C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex 24 Single Indexes for Illumina - Set #C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex 24 Single Indexes for Illumina - Set #D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<li>Compatible with<span> </span><a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24">D-Plex Small RNA-seq kit for Illumina</a> </li>
<li>Multiplexing up to <strong>96 samples</strong> when combining Set A, B, C and D</li>
<li>Allows for identification of index hopping</li>
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'meta_title' => 'D-Plex Unique Dual Indexes for Illumina - RNA Library Prep Kit - Set A | Diagenode',
'meta_keywords' => 'RNA kit; small RNA kit; RNA-seq kit; low input RNA-seq kit; small RNA-seq, template switching kit, RNA-seq library preparation, D-Plex, higher RNA diversity',
'meta_description' => 'Compatible with D-Plex RNA-seq kits - Unique Dual Indexes for Illumina - Index hopping mitigation - Suitable for ultra-low input - Contains UMI - Multiplexing up to 48 samples',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A">C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li>C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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'description' => '<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2 style="font-size: 22px;">DNA断片化、ライブラリー調製、自動化：NGSのワンストップショップ</h2>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-12 medium-12 large-12 columns">
<h4>1. 断片化装置を選択してください：150 bp〜75 kbの範囲でDNAを断片化します。</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-pico-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/bioruptor_pico.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/megaruptor2-1-unit"><img src="https://www.diagenode.com/img/product/shearing_technologies/B06010001_megaruptor2.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-one-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/br-one-profil.png" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns">5μlまで断片化：150 bp〜2 kb<br />NGS DNAライブラリー調製およびFFPE核酸抽出に最適で、</td>
<td class="small-4 medium-4 large-4 columns">2 kb〜75 kbの範囲をできます。<br />メイトペアライブラリー調製および長いフラグメントDNAシーケンシングに最適で、この軽量デスクトップデバイスで</td>
<td class="small-4 medium-4 large-4 columns">20または50μlの断片化が可能です。</td>
</tr>
</tbody>
</table>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-8 medium-8 large-8 columns">
<h4>2. 最適化されたライブラリー調整キットを選択してください。</h4>
</th>
<th class="small-4 medium-4 large-4 columns">
<h4>3. ライブラリー前処理自動化を選択して、比類のないデータ再現性を実感</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><img src="https://www.diagenode.com/img/product/kits/microPlex_library_preparation.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/ideal-library-preparation-kit-x24-incl-index-primer-set-1-24-rxns"><img src="https://www.diagenode.com/img/product/kits/box_kit.jpg" style="display: block; margin-left: auto; margin-right: auto;" height="173" width="250" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/sx-8g-ip-star-compact-automated-system-1-unit"><img src="https://www.diagenode.com/img/product/automation/B03000002%20_ipstar_compact.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">50pgの低入力：MicroPlex Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">5ng以上：iDeal Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">Achieve great NGS data easily</td>
</tr>
</tbody>
</table>
</div>
</div>
<blockquote>
<div class="row">
<div class="small-12 medium-12 large-12 columns"><span class="label" style="margin-bottom: 16px; margin-left: -22px; font-size: 15px;">DiagenodeがNGS研究にぴったりなプロバイダーである理由</span>
<p>Diagenodeは15年以上もエピジェネティクス研究に専念、専門としています。 ChIP研究クロマチン用のユニークな断片化システムの開発から始まり、 専門知識を活かし、5μlのせん断体積まで可能で、NGS DNAライブラリーの調製に最適な最先端DNA断片化装置の開発にたどり着きました。 我々は以来、ChIP-seq、Methyl-seq、NGSライブラリー調製用キットを研究開発し、業界をリードする免疫沈降研究と同様に、ライブラリー調製を自動化および完結させる独自の自動化システムを開発にも成功しました。</p>
<ul>
<li>信頼されるせん断装置</li>
<li>様々なインプットからのライブラリ作成キット</li>
<li>独自の自動化デバイス</li>
</ul>
</div>
</div>
</blockquote>
<div class="row">
<div class="small-12 columns">
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#panel1a">次世代シーケンシングへの理解とその専門知識</a>
<div id="panel1a" class="content">
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<p><strong>次世代シーケンシング (NGS)</strong> ）は、著しいスケールとハイスループットでシーケンシングを行い、1日に数十億もの塩基生成を可能にします。 NGSのハイスループットは迅速でありながら正確で、再現性のあるデータセットを実現し、さらにシーケンシング費用を削減します。 NGSは、ゲノムシーケンシング、ゲノム再シーケンシング、デノボシーケンシング、トランスクリプトームシーケンシング、その他にDNA-タンパク質相互作用の検出やエピゲノムなどを示します。 指数関数的に増加するシーケンシングデータの需要は、計算分析の障害や解釈、データストレージなどの課題を解決します。</p>
<p>アプリケーションおよび出発物質に応じて、数百万から数十億の鋳型DNA分子を大規模に並行してシーケンシングすることが可能です。その為に、異なる化学物質を使用するいくつかの市販のNGSプラットフォームを利用することができます。 NGSプラットフォームの種類によっては、事前準備とライブラリー作成が必要です。</p>
<p>NGSにとっても、特にデータ処理と分析に関した大きな課題はあります。第3世代技術はゲノミクス研究にさらに革命を起こすであろうと大きく期待されています。</p>
</div>
</div>
<div class="row">
<div class="small-6 medium-6 large-6 columns">
<p><strong>NGS アプリケーション</strong></p>
<ul>
<li>全ゲノム配列決定</li>
<li>デノボシーケンシング</li>
<li>標的配列</li>
<li>Exomeシーケンシング</li>
<li>トランスクリプトーム配列決定</li>
<li>ゲノム配列決定</li>
<li>ミトコンドリア配列決定</li>
<li>DNA-タンパク質相互作用（ChIP-seq</li>
<li>バリアント検出</li>
<li>ゲノム仕上げ</li>
</ul>
</div>
<div class="small-6 medium-6 large-6 columns">
<p><strong>研究分野におけるNGS:</strong></p>
<ul>
<li>腫瘍学</li>
<li>リプロダクティブ・ヘルス</li>
<li>法医学ゲノミクス</li>
<li>アグリゲノミックス</li>
<li>複雑な病気</li>
<li>微生物ゲノミクス</li>
<li>食品・環境ゲノミクス</li>
<li>創薬ゲノミクス - パーソナライズド・メディカル</li>
</ul>
</div>
<div class="small-12 medium-12 large-12 columns">
<p><strong>NGSの用語</strong></p>
<dl>
<dt>リード（読み取り）</dt>
<dd>この装置から得られた連続した単一のストレッチ</dd>
<dt>断片リード</dt>
<dd>フラグメントライブラリからの読み込み。 シーケンシングプラットフォームに応じて、読み取りは通常約100〜300bp。</dd>
<dt>断片ペアエンドリード</dt>
<dd>断片ライブラリーからDNA断片の各末端2つの読み取り。</dd>
<dt>メイトペアリード</dt>
<dd>大きなDNA断片（通常は予め定義されたサイズ範囲）の各末端から2つの読み取り。</dd>
<dt>カバレッジ（例）</dt>
<dd>30×適用範囲とは、参照ゲノム中の各塩基対が平均30回の読み取りを示す。</dd>
</dl>
</div>
</div>
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2>NGSプラットフォーム</h2>
<h3><a href="http://www.illumina.com" target="_blank">イルミナ</a></h3>
<p>イルミナは、クローン的に増幅された鋳型DNA（クラスター）上に位置する、蛍光標識された可逆的鎖ターミネーターヌクレオチドを用いた配列別合成技術を使用。 DNAクラスターは、ガラスフローセルの表面上に固定化され、 ワークフローは、4つのヌクレオチド（それぞれ異なる蛍光色素で標識された）の組み込み、4色イメージング、色素や末端基の切断、取り込み、イメージングなどを繰り返します。フローセルは大規模な並列配列決定を受ける。 この方法により、単一蛍光標識されたヌクレオチドの制御添加によるモノヌクレオチドのエラーを回避する可能性があります。 読み取りの長さは、通常約100〜150 bpです。</p>
<h3><a href="http://www.lifetechnologies.com" target="_blank">イオン トレント</a></h3>
<p>イオントレントは、半導体技術チップを用いて、合成中にヌクレオチドを取り込む際に放出されたプロトンを検出します。 これは、イオン球粒子と呼ばれるビーズの表面にエマルションPCR（emPCR）を使用し、リンクされた特定のアダプターを用いてDNA断片を増幅します。 各ビーズは1種類のDNA断片で覆われていて、異なるDNA断片を有するビーズは次いで、チップの陽子感知ウェル内に配置されます。 チップには一度に4つのヌクレオチドのうちの1つが浸水し、このプロセスは異なるヌクレオチドで15秒ごとに繰り返されます。 配列決定の間に4つの塩基の各々が1つずつ導入されます、組み込みの場合はプロトンが放出され、電圧信号が取り込みに比例して検出されます。.</p>
<h3><a href="http://www.pacificbiosciences.com" target="_blank">パシフィック バイオサイエンス</a></h3>
<p>パシフィックバイオサイエンスでは、20kbを超える塩基対の読み取りも、単一分子リアルタイム（SMRT）シーケンシングによる構造および細胞タイプの変化を観察することができます。 このプラットフォームでは、超長鎖二本鎖DNA（dsDNA）断片が、Megaruptor（登録商標）のようなDiagenode装置を用いたランダムシアリングまたは目的の標的領域の増幅によって生成されます。 SMRTbellライブラリーは、ユニバーサルヘアピンアダプターをDNA断片の各末端に連結することによって生成します。 サイズ選択条件による洗浄ステップの後、配列決定プライマーをSMRTbellテンプレートにアニーリングし、鋳型DNAに結合したDNAポリメラーゼを含む配列決定を、蛍光標識ヌクレオチドの存在下で開始。 各塩基が取り込まれると、異なる蛍光のパルスをリアルタイムで検出します。</p>
<h3><a href="https://nanoporetech.com" target="_blank">オックスフォード ナノポア</a></h3>
<p>Oxford Nanoporeは、単一のDNA分子配列決定に基づく技術を開発します。その技術により生物学的分子、すなわちDNAが一群の電気抵抗性高分子膜として位置するナノスケールの孔（ナノ細孔）またはその近くを通過し、イオン電流が変化します。 この変化に関する情報は、例えば4つのヌクレオチド（AまたはG r CまたはT）ならびに修飾されたヌクレオチドすべてを区別することによって分子情報に訳されます。 シーケンシングミニオンデバイスのフローセルは、数百個のナノポアチャネルのセンサアレイを含みます。 DNAサンプルは、Diagenode社のMegaruptor（登録商標）を用いてランダムシアリングによって生成され得る超長鎖DNAフラグメントが必要です。</p>
<h3><a href="http://www.lifetechnologies.com/be/en/home/life-science/sequencing/next-generation-sequencing/solid-next-generation-sequencing.html" target="_blank">SOLiD</a></h3>
<p>SOLiDは、ユニークな化学作用により、何千という個々のDNA分子の同時配列決定を可能にします。 それは、アダプター対ライブラリーのフラグメントが適切で、せん断されたゲノムDNAへのアダプターのライゲーションによるライブラリー作製から始まります。 次のステップでは、エマルジョンPCR（emPCR）を実施して、ビーズの表面上の個々の鋳型DNA分子をクローン的に増幅。 emPCRでは、個々の鋳型DNAをPCR試薬と混合し、水中油型エマルジョン内の疎水性シェルで囲まれた水性液滴内のプライマーコートビーズを、配列決定のためにロードするスライドガラスの表面にランダムに付着。 この技術は、シークエンシングプライマーへのライゲーションで競合する4つの蛍光標識されたジ塩基プローブのセットを使用します。</p>
<h3><a href="http://454.com/products/technology.asp" target="_blank">454</a></h3>
<p>454は、大規模並列パイロシーケンシングを利用しています。 始めに全ゲノムDNAまたは標的遺伝子断片の300〜800bp断片のライブラリー調製します。 次に、DNAフラグメントへのアダプターの付着および単一のDNA鎖の分離。 その後アダプターに連結されたDNAフラグメントをエマルジョンベースのクローン増幅（emPCR）で処理し、DNAライブラリーフラグメントをミクロンサイズのビーズ上に配置します。 各DNA結合ビーズを光ファイバーチップ上のウェルに入れ、器具に挿入します。 4つのDNAヌクレオチドは、配列決定操作中に固定された順序で連続して加えられ、並行して配列決定されます。</p>
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<p><span style="font-weight: 400;">Most of the major next-generation sequencing platforms require ligation of specific adaptor oligos to </span><a href="../applications/dna-rna-shearing"><span style="font-weight: 400;">fragmented DNA or RNA</span></a><span style="font-weight: 400;"> prior to sequencing</span></p>
<p><span style="font-weight: 400;">After input DNA has been fragmented, it is end-repaired and blunt-ended</span><span style="font-weight: 400;">. The next step is a A-tailing in which dAMP is added to the 3´ end of the blunt phosphorylated DNA fragments to prevent concatemerization and to allow the ligation of adaptors with complementary dT overhangs. In addition, barcoded adapters can be incorporated to facilitate multiplexing prior to or during amplification.</span></p>
<center><img src="https://www.diagenode.com/img/categories/library-prep/flux.png" /></center>
<p><span style="font-weight: 400;">Diagenode offers a comprehensive product portfolio for library preparation:<br /></span></p>
<strong><a href="https://www.diagenode.com/en/categories/Library-preparation-for-RNA-seq">D-Plex RNA-seq Library Preparation Kits</a></strong><br />
<p><span style="font-weight: 400;">Diagenode’s new RNA-sequencing solutions utilize the innovative c</span><span style="font-weight: 400;">apture and a</span><span style="font-weight: 400;">mplification by t</span><span style="font-weight: 400;">ailing and s</span><span style="font-weight: 400;">witching”</span><span style="font-weight: 400;">, a ligation-free method to produce DNA libraries for next generation sequencing from low input amounts of RNA. </span><span style="font-weight: 400;"></span><a href="../categories/Library-preparation-for-RNA-seq">Learn more</a></p>
<strong><a href="../categories/library-preparation-for-ChIP-seq">ChIP-seq and DNA sequencing library preparation solutions</a></strong><br />
<p><span style="font-weight: 400;">Our kits have been optimized for DNA library preparation used for next generation sequencing for a wide range of inputs. Using a simple three-step protocols, our</span><a href="http://www.diagenode.com/p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><span style="font-weight: 400;"> </span></a><span style="font-weight: 400;">kits are an optimal choice for library preparation from DNA inputs down to 50 pg. </span><a href="../categories/library-preparation-for-ChIP-seq">Learn more</a></p>
<a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span><strong>Bioruptor Pico - short fragments</strong></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">Our well-cited Bioruptor Pico is the shearing device of choice for chromatin and DNA fragmentation. Obtain uniform and tight fragment distributions between 150bp -2kb. </span><a href="../p/bioruptor-pico-sonication-device">Learn more</a></p>
<strong><a href="../p/megaruptor2-1-unit"><span href="../p/bioruptor-pico-sonication-device">Megaruptor</span>® - long fragments</a></strong><a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">The Megaruptor is designed to shear DNA from 3kb-75kb for long-read sequencing. <a href="../p/megaruptor2-1-unit">Learn more</a></span></p>
<span href="../p/bioruptor-pico-sonication-device"></span><span style="font-weight: 400;"></span></div>
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'name' => 'Low-nutrient diet in larvae stage causes enhancement in dopaminemodulation in adult brain due epigenetic imprinting.',
'authors' => 'Zúñiga-Hernández JM et al.',
'description' => '<p><span>Nutrient scarcity is a frequent adverse condition that organisms face during their development. This condition may lead to long-lasting effects on the metabolism and behaviour of adults due to developmental epigenetic modifications. Here, we show that reducing nutrient availability during larval development affects adult spontaneous activity and sleep behaviour, together with changes in gene expression and epigenetic marks in the mushroom bodies (MBs). We found that open chromatin regions map to 100 of 241 transcriptionally upregulated genes in the adult MBs, these new opening zones are preferentially located in regulatory zones such as promoter-TSS and introns. Importantly, opened chromatin at the Dopamine 1-like receptor 2 regulatory zones correlate with increased expression. In consequence, adult administration of a dopamine antagonist reverses increased spontaneous activity and diminished sleep time observed in response to early-life nutrient restriction. In comparison, reducing </span><i>dop1R2</i><span><span> </span>expression in MBs also ameliorates these effects, albeit to a lesser degree. These results lead to the conclusion that increased dopamine signalling in the MBs of flies reared in a poor nutritional environment underlies the behavioural changes observed due to this condition during development.</span></p>',
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'description' => '<p><span>Cobalt and Nickel show increasing pollution in our environment. We analyzed their combined toxicity and uptake mechanisms in the early food chain by investigation of bacteria and the ciliate Paramecium as a primary consumer. Exposing Paramecium to metals, we can dissect between phagocytotic uptake and phagocytosis independent uptake. Measurement of cellular content in bacteria after exposure shows a bias for Nickel when both are co-exposed and even increased levels in comparison to single exposure. Imitating the initial food chain, we fed these bacteria to paramecia measuring only phagocytotic uptake. The cellular content shows a similar ratio of Nickel and Cobalt as in food bacteria suggesting that bacteria can selectively pre-accumulate metals for introduction into the food-chain. Analyzing the transcriptomic response of paramecia to sublethal doses of Nickel and Cobalt, Gene ontogogy (GO)-analysis indicates common deregulated pathways, e.g. the ammonium-transmembrane transport and ubiquitine-associated protein degradation. Redox related genes show a massive de-regulation of gene expression indicating cellular adaptation to increased ROS stress. As this suggests that both metals can target also the same cellular pathways, this is in agreement with increased toxicity of both metals in joint applications. Our data reveals complex routes of individual metals to enter the food chain.</span></p>',
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'name' => 'Manganese-Induced Toxicity in : What Can We Learn from theTranscriptome?',
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'description' => '<p>Manganese (Mn) is an essential ubiquitous transition metal and, when occupationally or environmentally overexposed, a well-known risk factor for several neurological pathologies. However, the molecular mechanisms underlying Mn-induced neurotoxicity are largely unknown. In this study, addressing RNA-Seq analysis, bioavailability and survival assays, key pathways of transcriptional responses to Mn overexposure were investigated in the model organism (), providing insights into the Mn-induced cellular stress and damage response. Comparative transcriptome analyses identified a large number of differentially expressed genes (DEGs) in nematodes exposed to MnCl, and functional annotation suggested oxidative nucleotide damage, unfolded protein response and innate immunity as major damage response pathways. Additionally, a time-dependent increase in the transcriptional response after MnCl exposure was identified by means of increased numbers of DEGs, indicating a time-dependent response and activation of the stress responses in Mn neurotoxicity. The data provided here represent a powerful transcriptomic resource in the field of Mn toxicity, and therefore, this study provides a useful basis for further planning of targeted mechanistic studies of Mn-induced neurotoxicity that are urgently needed in the face of increasing industrially caused environmental pollution with Mn.</p>',
'date' => '2022-09-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/36142660',
'doi' => '10.3390/ijms231810748',
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'name' => 'A Chromosomal Duplication Encompassing Interleukin-33 Causes aNovel Hyper IgE Phenotype Characterized by EosinophilicEsophagitis and Generalized Autoimmunity.',
'authors' => 'Marwaha A.K. et al.',
'description' => '<p>Interleukin (IL)-33 is a member of the IL-1 cytokine family and is secreted by fibroblasts, endothelial cells, and epithelial cells in tissues (lung, skin, and gastrointestinal) that are exposed to the environment. 1 IL-33 is stored in the nucleus and environmental allergens trigger RIPK1-caspase 8 ripoptosome activation in epithelial cells to enable IL-33 maturation and release. 2 IL-33 signals via its receptor ST2, which is found on multiple immune cells, such as eosinophils, mast cells, T cells, macrophages, basophils, and type 2 innate lymphoid cells, which all promote type-2 innate immune reactions. 3 Therefore, IL-33 plays a central role in promoting allergic inflammatory responses in diseases such as eosinophilic esophagitis (EoE), allergy to food/inhalants, asthma, and atopic dermatitis. Genome-wide association data of these conditions consistently report polymorphisms in the IL-33 gene as a risk factor. 4 Thus, IL-33 has been proposed as a therapeutic target for these conditions. In animal models, IL-33 has been mechanistically linked with allergic, autoimmune, rheumatologic, and inflammatory bowel diseases. 5 However, monogenic human diseases caused by either gain-of-function or loss-of-function variants in the IL-33 gene have not been reported previously.</p>',
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'name' => 'Functional microRNA targetome undergoes degeneration-induced shift inthe retina.',
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'description' => '<p>BACKGROUND: MicroRNA (miRNA) play a significant role in the pathogenesis of complex neurodegenerative diseases including age-related macular degeneration (AMD), acting as post-transcriptional gene suppressors through their association with argonaute 2 (AGO2) - a key member of the RNA Induced Silencing Complex (RISC). Identifying the retinal miRNA/mRNA interactions in health and disease will provide important insight into the key pathways miRNA regulate in disease pathogenesis and may lead to potential therapeutic targets to mediate retinal degeneration. METHODS: To identify the active miRnome targetome interactions in the healthy and degenerating retina, AGO2 HITS-CLIP was performed using a rodent model of photoreceptor degeneration. Analysis of publicly available single-cell RNA sequencing (scRNAseq) data was performed to identify the cellular location of AGO2 and key members of the microRNA targetome in the retina. AGO2 findings were verified by in situ hybridization (RNA) and immunohistochemistry (protein). RESULTS: Analysis revealed a similar miRnome between healthy and damaged retinas, however, a shift in the active targetome was observed with an enrichment of miRNA involvement in inflammatory pathways. This shift was further demonstrated by a change in the seed binding regions of miR-124-3p, the most abundant retinal AGO2-bound miRNA, and has known roles in regulating retinal inflammation. Additionally, photoreceptor cluster miR-183/96/182 were all among the most highly abundant miRNA bound to AGO2. Following damage, AGO2 expression was localized to the inner retinal layers and more in the OLM than in healthy retinas, indicating a locational miRNA response to retinal damage. CONCLUSIONS: This study provides important insight into the alteration of miRNA regulatory activity that occurs as a response to retinal degeneration and explores the miRNA-mRNA targetome as a consequence of retinal degenerations. Further characterisation of these miRNA/mRNA interactions in the context of the degenerating retina may provide an important insight into the active role these miRNA may play in diseases such as AMD.</p>',
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'description' => '<div class="row"><div class="small-12 medium-4 large-4 columns"><div class="panel"><h3><em>lncRNAs in cancer</em></h3><p class="text-left">Gastrointestinal cancer: Gastrointestinal cancers occur as a result of dysregulated signaling pathways and cellular processes, such as the cell cycle or apoptosis. LncRNAs can regulate proliferation of gastrointestinal cancer through interacting with RNA targets, localization to chromatin, or binding to proteins. Read more about Long non-coding RNAs: crucial regulators of gastrointestinal cancer cell proliferation</p><h3><em>microRNAs in cancer</em></h3><p class="text-left">miR-155 has been found overexpressed in many cancer types including hematopoietic cancers, breast, lung and colon cancer<br /><br /> <a href="https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf">https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf</a></p></div></div><div class="small-12 medium-8 large-8 columns"><center><img src="https://www.diagenode.com/img/cancer/long-non-coding-rna.jpg" width="550" height="345" /></center><p></p><p>Various non-coding RNAs (ncRNAs) have been found to function as key regulators for transcription, chromatin remodelling, and post-transcriptional modification, thus making them relevant in oncogenesis, both as tumor suppressors and as drivers. For example, a number of microRNAs (miRNAs), one of the more well-studied types of ncRNAs, have been identified as potential cancer biomarkers and clinical targets. Other ncRNAs, such as long noncoding RNAs are involved in cancer regulation and proliferation. Understanding the role of ncRNAs will be critical for cancer research and therapeutic development.</p></div></div>',
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<h6 style="height:60px">D-Plex Unique Dual Indexes for Illumina - Set B</h6>
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
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<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
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<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
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'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li><span style="font-weight: 400;">An innovative technology with template switching and UMIs</span></li>
<li><span style="font-weight: 400;">Ultra-low input capability, down to 50 pg for total RNAs</span></li>
<li><span style="font-weight: 400;">Capture the widest possible diversity of RNAs for rich content</span></li>
<li><span>Get high sensitivity data even from difficult samples, such as degraded, FFPE samples</span></li>
<li><span style="font-weight: 400;">Enjoy a fast, easy, single tube protocol</span></li>
</ul>
</div>
<p></p>
<p></p>
<h4></h4>
<center><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/dplex-total-manual.pdf" target="_blank" title="dplex total rnaseq user manual"><strong>Download the manual</strong></a></center>
<p></p>
<p>D-Plex Total RNA-seq Library Preparation Kit is a tool designed for the study of the whole coding and non-coding transcriptome. <span>The kit is using the</span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank"><span> </span>D-Plex technology</a><span><span> </span>to generate directional libraries for Illumina sequencing directly from total RNAs, mRNAs that has already been enriched by poly(A) selection, or RNAs that has already been depleted of rRNAs.</span></p>
<p><span>The D-Plex technology utilizes two innovative ligation-free mechanisms - poly(A) tailing and template switching - to produce sequencing libraries from ultra-low input amounts, down to 50 pg for total RNAs, mRNAs or rRNA-depleted RNAs. Combined with unique molecular identifiers (UMI), this complete solution delivers a high-sensitivity detection method for comprehensive analysis of the transcriptome. It accurately measures gene and transcript abundance and captures the widest possible diversity of RNAs, including known and novel features in coding and non-coding RNAs.</span></p>
<p><span></span><span>D-Plex Total RNA-seq Kit offers a time saving protocol that can be completed within 5 hours and requires minimal hands-on time. <span>The library preparation takes place in a </span><span>single tube</span><span>, increasing the efficiency tremendously. This new solution has been extensively validated for both intact and highly degraded RNA samples, including that derived from FFPE preparations.</span></span></p>
<p><span><span>D-Plex Total RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Unique Dual Indexes were designed and validated to fit the D-Plex technology and are available separately:</span></span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C </a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<div class="extra-spaced" align="center"></div>
<div class="row">
<div class="carrousel" style="background-position: center;">
<div class="slick">
<div>
<h3>Greater sensitivity to detect novel transcripts</h3>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/CPM_totalrna.png" alt="total RNA kit" /></center></div>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq kit enables great transcript detection, even when starting from very low RNA inputs or working with challenging FFPE samples. Increased number of transcripts detected at 1x coverage is an indicator of greater sensitivity.</p>
</div>
</div>
<div>
<h3>Highest diversity to facilitate biomarker discovery</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_rdep_biotypes.png" alt="total RNA diversity" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq libraries capture efficiently all RNA biotypes present in the sample of interest, both coding and non-coding RNAs or small and long RNAs.</p>
</div>
</div>
<div>
<h3>Consistent expression profiling across a wide range of RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_and_umeg_rdep_violin_tpm_10.png" alt="total RNA" width="450px" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Gene expression profiling (including protein coding exons and introns) obtained with D-Plex Total RNA-seq kit is conserved for decreasing RNA inputs, keeping transcript diversity across the range of RNA inputs.</p>
</div>
</div>
<div>
<h3>Superior library complexity at low RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/robustness_corr_HUR_HuMEg_rrna.png" alt="total RNA" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Correlation analysis of the protein coding genes detected indicates superior transcript expression correlation between low and high RNA inputs. This result demonstrates that D-Plex is an ideal solution for challenging samples such as FFPE preparations.</p>
</div>
</div>
</div>
</div>
</div>',
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'label2' => 'Indexes',
'info2' => '<p><span>Specific D-Plex indexes </span><span>were designed and validated to fit the D-Plex technology for Illumina sequencing and </span><span>are not included in the kit. They can be bought separately according to your needs. Please choose the format that suits you best among the compatible references to:</span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex UDI Module - Set C" target="_blank"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<p></p>
<p>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</p>',
'label3' => 'Data analysis',
'info3' => '<p><span>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</span></p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="Small RNA seq Bioinformatics pipeline" width="925" height="196" /></p>',
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'slug' => 'D-Plex-Total-RNA-seq-Library-Prep-x24',
'meta_title' => 'D-Plex Total RNA-seq Library Prep Kit for Illumina | Diagenode ',
'meta_keywords' => 'RNA-seq kit, RNA-seq library preparation; low input RNA-seq, UMI RNA-seq, UDI RNA-seq, D-Plex, higher RNA sensitivity',
'meta_description' => 'RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (50 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with FFPE samples - User-friendly and fast protocol',
'modified' => '2023-11-16 09:09:25',
'created' => '2021-04-13 14:10:40',
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'id' => '3037',
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'name' => 'D-Plex Small RNA-seq Kit for Illumina',
'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li>Ultra-low input capability, down to 10 pg for small RNAs and 100 pg for total RNA samples</li>
<li>High library complexity providing novel and diverse transcript detection</li>
<li>Versatile integartion into numerpus transcriptomic analysis workflow: ribosome profiling, CLIP-seq, RIP-seq and other</li>
<li>Improved quantification accuracy through the use of UMIs</li>
<li>Easy to use with minimal hands-on time: one day, one tube protocol</li>
</ul>
</div>
<p></p>
<p></p>
<center><em><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/MA-D-Plex.pdf" target="_blank" title="D-Plex Small RNA library preparation user manual"><strong>Download the manual</strong></a></em></center>
<p>D-Plex Small RNA-seq Library Prep Kit is a tool designed for the study of the <strong>small non-coding transcriptome</strong>. The kit is using the<a href="https://www.diagenode.com/en/pages/dplex" target="_blank"> D-Plex technology</a> to generate small RNA libraries for Illumina sequencing directly from total RNAs or enriched small RNAs.</p>
<p>The D-Plex technology utilizes two innovative <strong>ligation-free mechanisms</strong> - <strong>poly(A) tailing</strong> and <strong>template switching</strong> - to produce sequencing libraries from ultra-low input amounts, down to 10 pg for small RNAs and 100 pg for total RNAs. Combined with <strong>unique molecular identifiers</strong> (UMI), this complete solution ensures a <strong>realistic</strong> and <strong>accurate representation of diverse small RNA species</strong> (such as microRNAs, piRNAs, tRNAs, and siRNAs) with minimized quantitative bias.</p>
<p>D-Plex Small RNA-seq Kit offers a time saving protocol that can be completed within <strong>5 hours</strong> and requires minimal hands-on time. <span>The library preparation takes place in a </span><strong>single tube</strong><span>, increasing the efficiency tremendously. </span></p>
<p>D-Plex Small RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Indexes were designed and validated to fit the D-Plex technology and are available separately:</p>
<p><strong>For D-Plex UDI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><strong>For D-Plex SI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-A" title="D-Plex SI Module - Set A" target="_blank">C05030010 - D-Plex Single Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-B" title="D-Plex SI Module - Set B" target="_blank">C05030011 - D-Plex Single Indexes for Illumina - Set B</a></li>
</ul>
<p><strong>D-Plex is also available for MGI sequencing, check <a href="https://www.diagenode.com/en/p/d-plex-small-RNA-dnbseq-kit-x24" target="_blank">here</a>!</strong></p>
<p></p>
<p></p>
<p></p>
<center><strong>The D-Plex smRNA-seq is a versatile tool for a plethora of transciptomic analysis</strong></center>
<p></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/versatile-circle.png" /></center>
<p></p>
<p>With low-input capability, accuracy and ability to incorporate different transcript classes independent of chemical moieties, the strong D-Plex library-preparation method can be included in various analysis workflows (i.e., ribosome profiling, genome-wide mapping of protein: RNA binding sites).</p>
<p></p>
<p></p>',
'label1' => 'Characteristics and library prep. workflow ',
'info1' => '<div class="row">
<div class="small-12 columns">
<p><strong>High library diversity for ultra-low inputs</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-diversity-fig1.png" /></center></div>
<div class="small-12 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig2-captures.png" /></center></div>
</div>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>D-Plex smRNA-seq captures the smRNA complexity of the sample in a sensitive manner. A large number of features is detected for each biotype at a CPM ≥5, as shown above for different species.</p>
</div>
</div>
<p></p>
<p><strong>Accurate representation of the smRNA content of a sample</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-ratplasma.png" /></center></div>
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-miRNAdistribution.png" /></center></div>
</div>
<div class="row">
<div class="small-6 columns">
<p><strong>Accurate identification of PCR duplicates in low-input samples using unique molecular identifiers (UMIs).</strong><br />The UMI read deduplication that is embedded in the D-Plex construct is used to avoid over-estimation of transcripts by identifying clones generated during the PCR amplification of the library. Despite limiting starting RNA input, the UMI correction removed technical copies of transcripts, maintaining the initial sample abundance.</p>
</div>
<div class="small-6 columns">
<p>D-Plex (template switching) technology enables recovery of the initial miRNA distribution of the miRXplore Universal Reference (Miltenyi Biotech Inc., Cat. No. 130-093-521). In contrast, ligation-based kits display a strong distortion of the miRNA equimolar distribution initially present in the pool, indicating sample incorporation bias.</p>
</div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-foldchange.jpg" /></center></div>
<div class="small-6 columns"><strong>D-Plex smRNA-seq in association with the MGcount analysis correlates strongly (R² > 0.8) with the RT-qPCR analysis, which is considered the gold-standard for accuracy. </strong><br />The figures shows a fold-change accuracy of a panel of 20 different small-RNA (figure taken from MGcount publication - Hita et al., BMC bioinformatics, 23-39(2022).</div>
</div>
<p></p>
<p></p>
<p><strong>Maximize information from the regulatory transcriptome using D-Plex small RNA-seq kit and MGcount*</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-multialignments.png" /></center></div>
<div class="small-6 columns">By analyzing D-Plex dataset with MGcount, more reads are kept during the analysis, which ultimately preserves biological complexity linked to ncRNA expression without decreasing the accuracy of detection. <br /><span style="line-height: 1em;"><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></span></div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>The D-Plex technology uses two innovative ligation-free mechanisms, <strong>poly(A) tailing and template switching</strong>, to produce sequencing libraries from ultra-low input amounts.</p>
</div>
</div>
<div class="row" style="background-color: #f5f5f5;">
<div class="small-12 columns"><br />
<p><strong>WORKFLOW</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/FL-Image-Workflow.jpg" width="50%" /></center></div>
<div class="small-12 columns">
<p style="text-align: justify; padding: 15px;"><br />RNA molecules are polyadenylated at the 3’-end and primed with an oligo(dT) primer containing part of the ILMN 3’-adapter sequence. The addition of a template-switching oligonucleotide during cDNA synthesis enables fusion of part of the ILMN 5’-adapter during reverse transcription. After PCR, the library comprises double stranded DNA constructs that are ready for sequencing.</p>
</div>
</div>
<p><br /><br /></p>
<p></p>
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'label2' => 'Data analysis',
'info2' => '<p>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
<p>Need help for your bioinformatics analysis of miRNA?</p>
<p></p>
<div class="small-12 medium-3 large-3 columns"><center><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"><img src="https://www.diagenode.com/img/product/kits/dplex/miRMaster_logo.png" /></a></center>
<p> </p>
<p> </p>
</div>
<div class="small-12 medium-9 large-9 columns">
<p>Diagenode has collaborated with <strong>Andreas Keller</strong> and <strong>Tobias Fehlmann</strong> to develop a new bioinformatics pipeline for <span>the <strong>prediction of novel miRNAs</strong>.</span><br /><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"></a></p>
<ul>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster2">miRMaster 2</a></li>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank">miRMaster</a></li>
</ul>
</div>
<p> </p>
<p> </p>',
'label3' => 'Data analysis - Counting using MGcount',
'info3' => '<p>The counting or expression level calculation is the last step of the processing to generate an expression level matrix. We recommend using MGcount*, a counting tool developed at Diagenode with a suitable annotations file that include the small RNA transcripts that are object of study. MGcount, built on top of featureCounts, employs a flexible quantification approach to deal with datasets containing multiple RNA biotypes such as D-plex libraries. Non-coding RNAs varying in length, biogenesis, and function, may overlap in a genomic region, and are sometimes present in the genome with a high copy number. Consequently, reads may align equally well to more than one position in the reference genome or/and align to a position where more than one annotated transcript is located. MGcount employs two strategies to quantify these reads. Firstly, it assigns reads in rounds prioritizing small-RNAs over long-RNAs ensuring the unbiased read attribution to intergenic and intragenic small RNAs while preventing host-genes transcription over-estimation. Secondly, MGcount collapses loci where reads consistently multi-map into communities of loci with a graph-based approach. The resultant communities are groups of biologically related loci with nearly identical sequence. Subsequently, quantification of reads is performed at the loci-community level, reducing multi-alignments ambiguity at individual locus. Ultimately, this strategy maximizes the transcriptome information analysed and improves the interrogation of non-coding RNAs. MGcount software repository provides annotation files for A. thaliana, H. sapiens, M. musculus and C. elegans. The required input files for MGcount are a .txt file listing the paths to the alignment input files (.bam format) and the annotations file (.gtf format). The output directory path has to be provided as an input as well.</p>
<p><strong>Example command:</strong></p>
<p style="background-color: #f0f0f0;">MGcount -T 2 --gtf Homo_sapiens.GRCh38.gtf --outdir outputs --bam_infiles input_bamfilenames.txt</p>
<p>MGcount provides the choice to enable/disable the quantification of all RNA biotypes included in the annotation file in the form of "communities" as optional parameters for small-RNA (--ml_flag_small) and long-RNA (--ml_flag_long). Both are enabled by default. The main output of MGcount is the count_matrix.csv file containing an expression matrix that can be imported to R or any other software for downstream analyses.<br />A full user guide for MGCount is available here: <a href="https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html">https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html</a>.</p>
<p>Other standard counting tools such as featureCounts or HTSeq-counts can also be used alternatively. Given the high complexity of D-Plex libraries, we recommend having a clear understanding of the scientific question and the goal of the project before proceeding to the choice of the counting method as this will strongly impact downstream analyses. Diagenode provides bioinformatics support as a service (please, contact us if you need help with data analysis).</p>
<p>Notice that D-Plex produces forward-stranded data. Stranded libraries have the benefit that reads map to the genome strand where they were originated from. Therefore, when estimating transcript expression, reads aligned to the forward strand should be assigned only to transcript features in the forward strand whereas reads aligned to the reverse strand should be assigned only to transcript features in the reverse strand. For this, make sure you select “stranded mode” in any tool of choice. Stranded mode is selected by default in MGcount.</p>
<p><em><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></em></p>',
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'meta_title' => 'D-Plex Small RNA-seq Library Prep Kit | Diagenode ',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set A includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
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<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex 24 Single Indexes for Illumina - Set #C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex 24 Single Indexes for Illumina - Set #D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
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<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A">C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li>C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
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<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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'name' => '次世代シーケンシング',
'description' => '<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2 style="font-size: 22px;">DNA断片化、ライブラリー調製、自動化：NGSのワンストップショップ</h2>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-12 medium-12 large-12 columns">
<h4>1. 断片化装置を選択してください：150 bp〜75 kbの範囲でDNAを断片化します。</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-pico-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/bioruptor_pico.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/megaruptor2-1-unit"><img src="https://www.diagenode.com/img/product/shearing_technologies/B06010001_megaruptor2.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-one-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/br-one-profil.png" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns">5μlまで断片化：150 bp〜2 kb<br />NGS DNAライブラリー調製およびFFPE核酸抽出に最適で、</td>
<td class="small-4 medium-4 large-4 columns">2 kb〜75 kbの範囲をできます。<br />メイトペアライブラリー調製および長いフラグメントDNAシーケンシングに最適で、この軽量デスクトップデバイスで</td>
<td class="small-4 medium-4 large-4 columns">20または50μlの断片化が可能です。</td>
</tr>
</tbody>
</table>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-8 medium-8 large-8 columns">
<h4>2. 最適化されたライブラリー調整キットを選択してください。</h4>
</th>
<th class="small-4 medium-4 large-4 columns">
<h4>3. ライブラリー前処理自動化を選択して、比類のないデータ再現性を実感</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><img src="https://www.diagenode.com/img/product/kits/microPlex_library_preparation.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/ideal-library-preparation-kit-x24-incl-index-primer-set-1-24-rxns"><img src="https://www.diagenode.com/img/product/kits/box_kit.jpg" style="display: block; margin-left: auto; margin-right: auto;" height="173" width="250" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/sx-8g-ip-star-compact-automated-system-1-unit"><img src="https://www.diagenode.com/img/product/automation/B03000002%20_ipstar_compact.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">50pgの低入力：MicroPlex Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">5ng以上：iDeal Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">Achieve great NGS data easily</td>
</tr>
</tbody>
</table>
</div>
</div>
<blockquote>
<div class="row">
<div class="small-12 medium-12 large-12 columns"><span class="label" style="margin-bottom: 16px; margin-left: -22px; font-size: 15px;">DiagenodeがNGS研究にぴったりなプロバイダーである理由</span>
<p>Diagenodeは15年以上もエピジェネティクス研究に専念、専門としています。 ChIP研究クロマチン用のユニークな断片化システムの開発から始まり、 専門知識を活かし、5μlのせん断体積まで可能で、NGS DNAライブラリーの調製に最適な最先端DNA断片化装置の開発にたどり着きました。 我々は以来、ChIP-seq、Methyl-seq、NGSライブラリー調製用キットを研究開発し、業界をリードする免疫沈降研究と同様に、ライブラリー調製を自動化および完結させる独自の自動化システムを開発にも成功しました。</p>
<ul>
<li>信頼されるせん断装置</li>
<li>様々なインプットからのライブラリ作成キット</li>
<li>独自の自動化デバイス</li>
</ul>
</div>
</div>
</blockquote>
<div class="row">
<div class="small-12 columns">
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#panel1a">次世代シーケンシングへの理解とその専門知識</a>
<div id="panel1a" class="content">
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<p><strong>次世代シーケンシング (NGS)</strong> ）は、著しいスケールとハイスループットでシーケンシングを行い、1日に数十億もの塩基生成を可能にします。 NGSのハイスループットは迅速でありながら正確で、再現性のあるデータセットを実現し、さらにシーケンシング費用を削減します。 NGSは、ゲノムシーケンシング、ゲノム再シーケンシング、デノボシーケンシング、トランスクリプトームシーケンシング、その他にDNA-タンパク質相互作用の検出やエピゲノムなどを示します。 指数関数的に増加するシーケンシングデータの需要は、計算分析の障害や解釈、データストレージなどの課題を解決します。</p>
<p>アプリケーションおよび出発物質に応じて、数百万から数十億の鋳型DNA分子を大規模に並行してシーケンシングすることが可能です。その為に、異なる化学物質を使用するいくつかの市販のNGSプラットフォームを利用することができます。 NGSプラットフォームの種類によっては、事前準備とライブラリー作成が必要です。</p>
<p>NGSにとっても、特にデータ処理と分析に関した大きな課題はあります。第3世代技術はゲノミクス研究にさらに革命を起こすであろうと大きく期待されています。</p>
</div>
</div>
<div class="row">
<div class="small-6 medium-6 large-6 columns">
<p><strong>NGS アプリケーション</strong></p>
<ul>
<li>全ゲノム配列決定</li>
<li>デノボシーケンシング</li>
<li>標的配列</li>
<li>Exomeシーケンシング</li>
<li>トランスクリプトーム配列決定</li>
<li>ゲノム配列決定</li>
<li>ミトコンドリア配列決定</li>
<li>DNA-タンパク質相互作用（ChIP-seq</li>
<li>バリアント検出</li>
<li>ゲノム仕上げ</li>
</ul>
</div>
<div class="small-6 medium-6 large-6 columns">
<p><strong>研究分野におけるNGS:</strong></p>
<ul>
<li>腫瘍学</li>
<li>リプロダクティブ・ヘルス</li>
<li>法医学ゲノミクス</li>
<li>アグリゲノミックス</li>
<li>複雑な病気</li>
<li>微生物ゲノミクス</li>
<li>食品・環境ゲノミクス</li>
<li>創薬ゲノミクス - パーソナライズド・メディカル</li>
</ul>
</div>
<div class="small-12 medium-12 large-12 columns">
<p><strong>NGSの用語</strong></p>
<dl>
<dt>リード（読み取り）</dt>
<dd>この装置から得られた連続した単一のストレッチ</dd>
<dt>断片リード</dt>
<dd>フラグメントライブラリからの読み込み。 シーケンシングプラットフォームに応じて、読み取りは通常約100〜300bp。</dd>
<dt>断片ペアエンドリード</dt>
<dd>断片ライブラリーからDNA断片の各末端2つの読み取り。</dd>
<dt>メイトペアリード</dt>
<dd>大きなDNA断片（通常は予め定義されたサイズ範囲）の各末端から2つの読み取り。</dd>
<dt>カバレッジ（例）</dt>
<dd>30×適用範囲とは、参照ゲノム中の各塩基対が平均30回の読み取りを示す。</dd>
</dl>
</div>
</div>
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2>NGSプラットフォーム</h2>
<h3><a href="http://www.illumina.com" target="_blank">イルミナ</a></h3>
<p>イルミナは、クローン的に増幅された鋳型DNA（クラスター）上に位置する、蛍光標識された可逆的鎖ターミネーターヌクレオチドを用いた配列別合成技術を使用。 DNAクラスターは、ガラスフローセルの表面上に固定化され、 ワークフローは、4つのヌクレオチド（それぞれ異なる蛍光色素で標識された）の組み込み、4色イメージング、色素や末端基の切断、取り込み、イメージングなどを繰り返します。フローセルは大規模な並列配列決定を受ける。 この方法により、単一蛍光標識されたヌクレオチドの制御添加によるモノヌクレオチドのエラーを回避する可能性があります。 読み取りの長さは、通常約100〜150 bpです。</p>
<h3><a href="http://www.lifetechnologies.com" target="_blank">イオン トレント</a></h3>
<p>イオントレントは、半導体技術チップを用いて、合成中にヌクレオチドを取り込む際に放出されたプロトンを検出します。 これは、イオン球粒子と呼ばれるビーズの表面にエマルションPCR（emPCR）を使用し、リンクされた特定のアダプターを用いてDNA断片を増幅します。 各ビーズは1種類のDNA断片で覆われていて、異なるDNA断片を有するビーズは次いで、チップの陽子感知ウェル内に配置されます。 チップには一度に4つのヌクレオチドのうちの1つが浸水し、このプロセスは異なるヌクレオチドで15秒ごとに繰り返されます。 配列決定の間に4つの塩基の各々が1つずつ導入されます、組み込みの場合はプロトンが放出され、電圧信号が取り込みに比例して検出されます。.</p>
<h3><a href="http://www.pacificbiosciences.com" target="_blank">パシフィック バイオサイエンス</a></h3>
<p>パシフィックバイオサイエンスでは、20kbを超える塩基対の読み取りも、単一分子リアルタイム（SMRT）シーケンシングによる構造および細胞タイプの変化を観察することができます。 このプラットフォームでは、超長鎖二本鎖DNA（dsDNA）断片が、Megaruptor（登録商標）のようなDiagenode装置を用いたランダムシアリングまたは目的の標的領域の増幅によって生成されます。 SMRTbellライブラリーは、ユニバーサルヘアピンアダプターをDNA断片の各末端に連結することによって生成します。 サイズ選択条件による洗浄ステップの後、配列決定プライマーをSMRTbellテンプレートにアニーリングし、鋳型DNAに結合したDNAポリメラーゼを含む配列決定を、蛍光標識ヌクレオチドの存在下で開始。 各塩基が取り込まれると、異なる蛍光のパルスをリアルタイムで検出します。</p>
<h3><a href="https://nanoporetech.com" target="_blank">オックスフォード ナノポア</a></h3>
<p>Oxford Nanoporeは、単一のDNA分子配列決定に基づく技術を開発します。その技術により生物学的分子、すなわちDNAが一群の電気抵抗性高分子膜として位置するナノスケールの孔（ナノ細孔）またはその近くを通過し、イオン電流が変化します。 この変化に関する情報は、例えば4つのヌクレオチド（AまたはG r CまたはT）ならびに修飾されたヌクレオチドすべてを区別することによって分子情報に訳されます。 シーケンシングミニオンデバイスのフローセルは、数百個のナノポアチャネルのセンサアレイを含みます。 DNAサンプルは、Diagenode社のMegaruptor（登録商標）を用いてランダムシアリングによって生成され得る超長鎖DNAフラグメントが必要です。</p>
<h3><a href="http://www.lifetechnologies.com/be/en/home/life-science/sequencing/next-generation-sequencing/solid-next-generation-sequencing.html" target="_blank">SOLiD</a></h3>
<p>SOLiDは、ユニークな化学作用により、何千という個々のDNA分子の同時配列決定を可能にします。 それは、アダプター対ライブラリーのフラグメントが適切で、せん断されたゲノムDNAへのアダプターのライゲーションによるライブラリー作製から始まります。 次のステップでは、エマルジョンPCR（emPCR）を実施して、ビーズの表面上の個々の鋳型DNA分子をクローン的に増幅。 emPCRでは、個々の鋳型DNAをPCR試薬と混合し、水中油型エマルジョン内の疎水性シェルで囲まれた水性液滴内のプライマーコートビーズを、配列決定のためにロードするスライドガラスの表面にランダムに付着。 この技術は、シークエンシングプライマーへのライゲーションで競合する4つの蛍光標識されたジ塩基プローブのセットを使用します。</p>
<h3><a href="http://454.com/products/technology.asp" target="_blank">454</a></h3>
<p>454は、大規模並列パイロシーケンシングを利用しています。 始めに全ゲノムDNAまたは標的遺伝子断片の300〜800bp断片のライブラリー調製します。 次に、DNAフラグメントへのアダプターの付着および単一のDNA鎖の分離。 その後アダプターに連結されたDNAフラグメントをエマルジョンベースのクローン増幅（emPCR）で処理し、DNAライブラリーフラグメントをミクロンサイズのビーズ上に配置します。 各DNA結合ビーズを光ファイバーチップ上のウェルに入れ、器具に挿入します。 4つのDNAヌクレオチドは、配列決定操作中に固定された順序で連続して加えられ、並行して配列決定されます。</p>
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<p><span style="font-weight: 400;">Most of the major next-generation sequencing platforms require ligation of specific adaptor oligos to </span><a href="../applications/dna-rna-shearing"><span style="font-weight: 400;">fragmented DNA or RNA</span></a><span style="font-weight: 400;"> prior to sequencing</span></p>
<p><span style="font-weight: 400;">After input DNA has been fragmented, it is end-repaired and blunt-ended</span><span style="font-weight: 400;">. The next step is a A-tailing in which dAMP is added to the 3´ end of the blunt phosphorylated DNA fragments to prevent concatemerization and to allow the ligation of adaptors with complementary dT overhangs. In addition, barcoded adapters can be incorporated to facilitate multiplexing prior to or during amplification.</span></p>
<center><img src="https://www.diagenode.com/img/categories/library-prep/flux.png" /></center>
<p><span style="font-weight: 400;">Diagenode offers a comprehensive product portfolio for library preparation:<br /></span></p>
<strong><a href="https://www.diagenode.com/en/categories/Library-preparation-for-RNA-seq">D-Plex RNA-seq Library Preparation Kits</a></strong><br />
<p><span style="font-weight: 400;">Diagenode’s new RNA-sequencing solutions utilize the innovative c</span><span style="font-weight: 400;">apture and a</span><span style="font-weight: 400;">mplification by t</span><span style="font-weight: 400;">ailing and s</span><span style="font-weight: 400;">witching”</span><span style="font-weight: 400;">, a ligation-free method to produce DNA libraries for next generation sequencing from low input amounts of RNA. </span><span style="font-weight: 400;"></span><a href="../categories/Library-preparation-for-RNA-seq">Learn more</a></p>
<strong><a href="../categories/library-preparation-for-ChIP-seq">ChIP-seq and DNA sequencing library preparation solutions</a></strong><br />
<p><span style="font-weight: 400;">Our kits have been optimized for DNA library preparation used for next generation sequencing for a wide range of inputs. Using a simple three-step protocols, our</span><a href="http://www.diagenode.com/p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><span style="font-weight: 400;"> </span></a><span style="font-weight: 400;">kits are an optimal choice for library preparation from DNA inputs down to 50 pg. </span><a href="../categories/library-preparation-for-ChIP-seq">Learn more</a></p>
<a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span><strong>Bioruptor Pico - short fragments</strong></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">Our well-cited Bioruptor Pico is the shearing device of choice for chromatin and DNA fragmentation. Obtain uniform and tight fragment distributions between 150bp -2kb. </span><a href="../p/bioruptor-pico-sonication-device">Learn more</a></p>
<strong><a href="../p/megaruptor2-1-unit"><span href="../p/bioruptor-pico-sonication-device">Megaruptor</span>® - long fragments</a></strong><a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">The Megaruptor is designed to shear DNA from 3kb-75kb for long-read sequencing. <a href="../p/megaruptor2-1-unit">Learn more</a></span></p>
<span href="../p/bioruptor-pico-sonication-device"></span><span style="font-weight: 400;"></span></div>
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'description' => '<p>Manganese (Mn) is an essential ubiquitous transition metal and, when occupationally or environmentally overexposed, a well-known risk factor for several neurological pathologies. However, the molecular mechanisms underlying Mn-induced neurotoxicity are largely unknown. In this study, addressing RNA-Seq analysis, bioavailability and survival assays, key pathways of transcriptional responses to Mn overexposure were investigated in the model organism (), providing insights into the Mn-induced cellular stress and damage response. Comparative transcriptome analyses identified a large number of differentially expressed genes (DEGs) in nematodes exposed to MnCl, and functional annotation suggested oxidative nucleotide damage, unfolded protein response and innate immunity as major damage response pathways. Additionally, a time-dependent increase in the transcriptional response after MnCl exposure was identified by means of increased numbers of DEGs, indicating a time-dependent response and activation of the stress responses in Mn neurotoxicity. The data provided here represent a powerful transcriptomic resource in the field of Mn toxicity, and therefore, this study provides a useful basis for further planning of targeted mechanistic studies of Mn-induced neurotoxicity that are urgently needed in the face of increasing industrially caused environmental pollution with Mn.</p>',
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'description' => '<p>Interleukin (IL)-33 is a member of the IL-1 cytokine family and is secreted by fibroblasts, endothelial cells, and epithelial cells in tissues (lung, skin, and gastrointestinal) that are exposed to the environment. 1 IL-33 is stored in the nucleus and environmental allergens trigger RIPK1-caspase 8 ripoptosome activation in epithelial cells to enable IL-33 maturation and release. 2 IL-33 signals via its receptor ST2, which is found on multiple immune cells, such as eosinophils, mast cells, T cells, macrophages, basophils, and type 2 innate lymphoid cells, which all promote type-2 innate immune reactions. 3 Therefore, IL-33 plays a central role in promoting allergic inflammatory responses in diseases such as eosinophilic esophagitis (EoE), allergy to food/inhalants, asthma, and atopic dermatitis. Genome-wide association data of these conditions consistently report polymorphisms in the IL-33 gene as a risk factor. 4 Thus, IL-33 has been proposed as a therapeutic target for these conditions. In animal models, IL-33 has been mechanistically linked with allergic, autoimmune, rheumatologic, and inflammatory bowel diseases. 5 However, monogenic human diseases caused by either gain-of-function or loss-of-function variants in the IL-33 gene have not been reported previously.</p>',
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'description' => '<p>BACKGROUND: MicroRNA (miRNA) play a significant role in the pathogenesis of complex neurodegenerative diseases including age-related macular degeneration (AMD), acting as post-transcriptional gene suppressors through their association with argonaute 2 (AGO2) - a key member of the RNA Induced Silencing Complex (RISC). Identifying the retinal miRNA/mRNA interactions in health and disease will provide important insight into the key pathways miRNA regulate in disease pathogenesis and may lead to potential therapeutic targets to mediate retinal degeneration. METHODS: To identify the active miRnome targetome interactions in the healthy and degenerating retina, AGO2 HITS-CLIP was performed using a rodent model of photoreceptor degeneration. Analysis of publicly available single-cell RNA sequencing (scRNAseq) data was performed to identify the cellular location of AGO2 and key members of the microRNA targetome in the retina. AGO2 findings were verified by in situ hybridization (RNA) and immunohistochemistry (protein). RESULTS: Analysis revealed a similar miRnome between healthy and damaged retinas, however, a shift in the active targetome was observed with an enrichment of miRNA involvement in inflammatory pathways. This shift was further demonstrated by a change in the seed binding regions of miR-124-3p, the most abundant retinal AGO2-bound miRNA, and has known roles in regulating retinal inflammation. Additionally, photoreceptor cluster miR-183/96/182 were all among the most highly abundant miRNA bound to AGO2. Following damage, AGO2 expression was localized to the inner retinal layers and more in the OLM than in healthy retinas, indicating a locational miRNA response to retinal damage. CONCLUSIONS: This study provides important insight into the alteration of miRNA regulatory activity that occurs as a response to retinal degeneration and explores the miRNA-mRNA targetome as a consequence of retinal degenerations. Further characterisation of these miRNA/mRNA interactions in the context of the degenerating retina may provide an important insight into the active role these miRNA may play in diseases such as AMD.</p>',
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'description' => '<h4>(Coming soon)</h4><p>D-Plex mRNA-seq Library Preparation Kit is a tool designed for the study of the whole coding transcriptome. <span>The kit is using the</span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank"><span> </span>D-Plex technology</a><span><span> </span>to generate directionnal libraries for Illumina sequencing directly from total RNAs or mRNAs that has already been enriched by poly(A) selection.</span></p><p><span>The D-Plex technology utilizes the innovative capture and amplification by tailing and switching, a ligation-free method for RNA library preparation from ultra-low input amounts. Combined with unique molecular identifiers (UMI), this complete solution delivers a high-sensitivity detection method for low-abundance or rare transcripts.</span></p><p><span></span><span>D-Plex mRNA-seq Kit offers a time saving protocol that can be completed within 5 hours and requires minimal hands-on time. <span>The library preparation takes place in a </span><span>single tube</span><span>, increasing the efficiency tremendously. This new solution has been extensively validated for both intact and highly degraded RNA samples, including that derived from FFPE preparations.</span></span></p><p><span><span>D-Plex Total RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Unique Dual Indexes were designed and validated to fit the D-Plex technology and are available separately:</span></span></p><ul><li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li><li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li></ul>',
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'info2' => '<p><span>Specific D-Plex indexes </span><span>were designed and validated to fit the D-Plex technology for Illumina sequencing and </span><span>are not included in the kit. They can be bought separately according to your needs. Please choose the format that suits you best among the compatible references to:</span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
</ul>
<p><span></span></p>
<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
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<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
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'meta_title' => 'D-Plex mRNA-seq Library Prep Kit | Diagenode ',
'meta_keywords' => 'RNA kit, RNA-seq kit, RNA-seq library preparation; low input RNA-seq, UMI RNA-seq, UDI RNA-seq, D-Plex, higher RNA sensitivity, Gene expression analysis',
'meta_description' => 'Gene expression analysis - Unique D-Plex technology for Illumina sequencing - Optimized for ultra-low input (50 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - User-friendly and fast protocol',
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'name' => 'D-Plex Total RNA-seq Kit for Illumina',
'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li><span style="font-weight: 400;">An innovative technology with template switching and UMIs</span></li>
<li><span style="font-weight: 400;">Ultra-low input capability, down to 50 pg for total RNAs</span></li>
<li><span style="font-weight: 400;">Capture the widest possible diversity of RNAs for rich content</span></li>
<li><span>Get high sensitivity data even from difficult samples, such as degraded, FFPE samples</span></li>
<li><span style="font-weight: 400;">Enjoy a fast, easy, single tube protocol</span></li>
</ul>
</div>
<p></p>
<p></p>
<h4></h4>
<center><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/dplex-total-manual.pdf" target="_blank" title="dplex total rnaseq user manual"><strong>Download the manual</strong></a></center>
<p></p>
<p>D-Plex Total RNA-seq Library Preparation Kit is a tool designed for the study of the whole coding and non-coding transcriptome. <span>The kit is using the</span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank"><span> </span>D-Plex technology</a><span><span> </span>to generate directional libraries for Illumina sequencing directly from total RNAs, mRNAs that has already been enriched by poly(A) selection, or RNAs that has already been depleted of rRNAs.</span></p>
<p><span>The D-Plex technology utilizes two innovative ligation-free mechanisms - poly(A) tailing and template switching - to produce sequencing libraries from ultra-low input amounts, down to 50 pg for total RNAs, mRNAs or rRNA-depleted RNAs. Combined with unique molecular identifiers (UMI), this complete solution delivers a high-sensitivity detection method for comprehensive analysis of the transcriptome. It accurately measures gene and transcript abundance and captures the widest possible diversity of RNAs, including known and novel features in coding and non-coding RNAs.</span></p>
<p><span></span><span>D-Plex Total RNA-seq Kit offers a time saving protocol that can be completed within 5 hours and requires minimal hands-on time. <span>The library preparation takes place in a </span><span>single tube</span><span>, increasing the efficiency tremendously. This new solution has been extensively validated for both intact and highly degraded RNA samples, including that derived from FFPE preparations.</span></span></p>
<p><span><span>D-Plex Total RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Unique Dual Indexes were designed and validated to fit the D-Plex technology and are available separately:</span></span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C </a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<div class="extra-spaced" align="center"></div>
<div class="row">
<div class="carrousel" style="background-position: center;">
<div class="slick">
<div>
<h3>Greater sensitivity to detect novel transcripts</h3>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/CPM_totalrna.png" alt="total RNA kit" /></center></div>
<div class="large-12 small-12 medium-12 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq kit enables great transcript detection, even when starting from very low RNA inputs or working with challenging FFPE samples. Increased number of transcripts detected at 1x coverage is an indicator of greater sensitivity.</p>
</div>
</div>
<div>
<h3>Highest diversity to facilitate biomarker discovery</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_rdep_biotypes.png" alt="total RNA diversity" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>D-Plex Total RNA-seq libraries capture efficiently all RNA biotypes present in the sample of interest, both coding and non-coding RNAs or small and long RNAs.</p>
</div>
</div>
<div>
<h3>Consistent expression profiling across a wide range of RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/HUR_and_umeg_rdep_violin_tpm_10.png" alt="total RNA" width="450px" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Gene expression profiling (including protein coding exons and introns) obtained with D-Plex Total RNA-seq kit is conserved for decreasing RNA inputs, keeping transcript diversity across the range of RNA inputs.</p>
</div>
</div>
<div>
<h3>Superior library complexity at low RNA inputs</h3>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/robustness_corr_HUR_HuMEg_rrna.png" alt="total RNA" /></center></div>
<div class="large-10 small-12 medium-10 large-centered medium-centered small-centered columns">
<p></p>
<p>Correlation analysis of the protein coding genes detected indicates superior transcript expression correlation between low and high RNA inputs. This result demonstrates that D-Plex is an ideal solution for challenging samples such as FFPE preparations.</p>
</div>
</div>
</div>
</div>
</div>',
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'info2' => '<p><span>Specific D-Plex indexes </span><span>were designed and validated to fit the D-Plex technology for Illumina sequencing and </span><span>are not included in the kit. They can be bought separately according to your needs. Please choose the format that suits you best among the compatible references to:</span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex UDI Module - Set C" target="_blank"><span>C05030023</span> - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex UDI Module - Set D" target="_blank"><span>C05030024</span> - D-Plex Unique Dual Indexes for Illumina - Set D </a></li>
</ul>
<p></p>
<p>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</p>',
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'info3' => '<p><span>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</span></p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="Small RNA seq Bioinformatics pipeline" width="925" height="196" /></p>',
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'slug' => 'D-Plex-Total-RNA-seq-Library-Prep-x24',
'meta_title' => 'D-Plex Total RNA-seq Library Prep Kit for Illumina | Diagenode ',
'meta_keywords' => 'RNA-seq kit, RNA-seq library preparation; low input RNA-seq, UMI RNA-seq, UDI RNA-seq, D-Plex, higher RNA sensitivity',
'meta_description' => 'RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (50 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with FFPE samples - User-friendly and fast protocol',
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'name' => 'D-Plex Small RNA-seq Kit for Illumina',
'description' => '<div class="small-12 medium-12 large-12 columns" style="border: 3px solid #B02736; padding: 10px; margin: 10px;">
<h2>Product Features</h2>
<ul style="list-style-type: disc;">
<li>Ultra-low input capability, down to 10 pg for small RNAs and 100 pg for total RNA samples</li>
<li>High library complexity providing novel and diverse transcript detection</li>
<li>Versatile integartion into numerpus transcriptomic analysis workflow: ribosome profiling, CLIP-seq, RIP-seq and other</li>
<li>Improved quantification accuracy through the use of UMIs</li>
<li>Easy to use with minimal hands-on time: one day, one tube protocol</li>
</ul>
</div>
<p></p>
<p></p>
<center><em><a class="chip diahome button" href="https://www.diagenode.com/files/products/kits/MA-D-Plex.pdf" target="_blank" title="D-Plex Small RNA library preparation user manual"><strong>Download the manual</strong></a></em></center>
<p>D-Plex Small RNA-seq Library Prep Kit is a tool designed for the study of the <strong>small non-coding transcriptome</strong>. The kit is using the<a href="https://www.diagenode.com/en/pages/dplex" target="_blank"> D-Plex technology</a> to generate small RNA libraries for Illumina sequencing directly from total RNAs or enriched small RNAs.</p>
<p>The D-Plex technology utilizes two innovative <strong>ligation-free mechanisms</strong> - <strong>poly(A) tailing</strong> and <strong>template switching</strong> - to produce sequencing libraries from ultra-low input amounts, down to 10 pg for small RNAs and 100 pg for total RNAs. Combined with <strong>unique molecular identifiers</strong> (UMI), this complete solution ensures a <strong>realistic</strong> and <strong>accurate representation of diverse small RNA species</strong> (such as microRNAs, piRNAs, tRNAs, and siRNAs) with minimized quantitative bias.</p>
<p>D-Plex Small RNA-seq Kit offers a time saving protocol that can be completed within <strong>5 hours</strong> and requires minimal hands-on time. <span>The library preparation takes place in a </span><strong>single tube</strong><span>, increasing the efficiency tremendously. </span></p>
<p>D-Plex Small RNA-seq Kit includes all buffers and enzymes necessary for the library preparation. Specific D-Plex Indexes were designed and validated to fit the D-Plex technology and are available separately:</p>
<p><strong>For D-Plex UDI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A" title="D-Plex UDI Module - Set A" target="_blank"><span>C05030021</span> - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex UDI Module - Set B" target="_blank"><span>C05030022</span> - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><strong>For D-Plex SI library construction:</strong></p>
<ul style="list-style-type: disc;">
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-A" title="D-Plex SI Module - Set A" target="_blank">C05030010 - D-Plex Single Indexes for Illumina - Set A</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Single-Indexes-Set-B" title="D-Plex SI Module - Set B" target="_blank">C05030011 - D-Plex Single Indexes for Illumina - Set B</a></li>
</ul>
<p><strong>D-Plex is also available for MGI sequencing, check <a href="https://www.diagenode.com/en/p/d-plex-small-RNA-dnbseq-kit-x24" target="_blank">here</a>!</strong></p>
<p></p>
<p></p>
<p></p>
<center><strong>The D-Plex smRNA-seq is a versatile tool for a plethora of transciptomic analysis</strong></center>
<p></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/versatile-circle.png" /></center>
<p></p>
<p>With low-input capability, accuracy and ability to incorporate different transcript classes independent of chemical moieties, the strong D-Plex library-preparation method can be included in various analysis workflows (i.e., ribosome profiling, genome-wide mapping of protein: RNA binding sites).</p>
<p></p>
<p></p>',
'label1' => 'Characteristics and library prep. workflow ',
'info1' => '<div class="row">
<div class="small-12 columns">
<p><strong>High library diversity for ultra-low inputs</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-diversity-fig1.png" /></center></div>
<div class="small-12 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig2-captures.png" /></center></div>
</div>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>D-Plex smRNA-seq captures the smRNA complexity of the sample in a sensitive manner. A large number of features is detected for each biotype at a CPM ≥5, as shown above for different species.</p>
</div>
</div>
<p></p>
<p><strong>Accurate representation of the smRNA content of a sample</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-ratplasma.png" /></center></div>
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-miRNAdistribution.png" /></center></div>
</div>
<div class="row">
<div class="small-6 columns">
<p><strong>Accurate identification of PCR duplicates in low-input samples using unique molecular identifiers (UMIs).</strong><br />The UMI read deduplication that is embedded in the D-Plex construct is used to avoid over-estimation of transcripts by identifying clones generated during the PCR amplification of the library. Despite limiting starting RNA input, the UMI correction removed technical copies of transcripts, maintaining the initial sample abundance.</p>
</div>
<div class="small-6 columns">
<p>D-Plex (template switching) technology enables recovery of the initial miRNA distribution of the miRXplore Universal Reference (Miltenyi Biotech Inc., Cat. No. 130-093-521). In contrast, ligation-based kits display a strong distortion of the miRNA equimolar distribution initially present in the pool, indicating sample incorporation bias.</p>
</div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-foldchange.jpg" /></center></div>
<div class="small-6 columns"><strong>D-Plex smRNA-seq in association with the MGcount analysis correlates strongly (R² > 0.8) with the RT-qPCR analysis, which is considered the gold-standard for accuracy. </strong><br />The figures shows a fold-change accuracy of a panel of 20 different small-RNA (figure taken from MGcount publication - Hita et al., BMC bioinformatics, 23-39(2022).</div>
</div>
<p></p>
<p></p>
<p><strong>Maximize information from the regulatory transcriptome using D-Plex small RNA-seq kit and MGcount*</strong></p>
<div class="row">
<div class="small-6 columns"><center><img src="https://www.diagenode.com/img/product/kits/dplex/dplex-fig-multialignments.png" /></center></div>
<div class="small-6 columns">By analyzing D-Plex dataset with MGcount, more reads are kept during the analysis, which ultimately preserves biological complexity linked to ncRNA expression without decreasing the accuracy of detection. <br /><span style="line-height: 1em;"><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></span></div>
</div>
<p></p>
<p></p>
<div class="row">
<div class="small-12 columns">
<p>The D-Plex technology uses two innovative ligation-free mechanisms, <strong>poly(A) tailing and template switching</strong>, to produce sequencing libraries from ultra-low input amounts.</p>
</div>
</div>
<div class="row" style="background-color: #f5f5f5;">
<div class="small-12 columns"><br />
<p><strong>WORKFLOW</strong></p>
<center><img src="https://www.diagenode.com/img/product/kits/dplex/FL-Image-Workflow.jpg" width="50%" /></center></div>
<div class="small-12 columns">
<p style="text-align: justify; padding: 15px;"><br />RNA molecules are polyadenylated at the 3’-end and primed with an oligo(dT) primer containing part of the ILMN 3’-adapter sequence. The addition of a template-switching oligonucleotide during cDNA synthesis enables fusion of part of the ILMN 5’-adapter during reverse transcription. After PCR, the library comprises double stranded DNA constructs that are ready for sequencing.</p>
</div>
</div>
<p><br /><br /></p>
<p></p>
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'info2' => '<p>A specific bioinformatics pipeline has been developed to process the special sequences present in the D-Plex construct, namely the UMI, the A-tail, and the template switch motif. All guidelines and free softwares are shared in the user manual. Subject to the compatibility of D-Plex constructs, other specific pipelines can be used.</p>
<p><img src="https://www.diagenode.com/img/product/kits/dplex/bioinfoPipe.png" alt="small RNA sequencing bioinformatics pipeline" caption="false" width="925" height="196" /></p>
<p>Need help for your bioinformatics analysis of miRNA?</p>
<p></p>
<div class="small-12 medium-3 large-3 columns"><center><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"><img src="https://www.diagenode.com/img/product/kits/dplex/miRMaster_logo.png" /></a></center>
<p> </p>
<p> </p>
</div>
<div class="small-12 medium-9 large-9 columns">
<p>Diagenode has collaborated with <strong>Andreas Keller</strong> and <strong>Tobias Fehlmann</strong> to develop a new bioinformatics pipeline for <span>the <strong>prediction of novel miRNAs</strong>.</span><br /><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank"></a></p>
<ul>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster2">miRMaster 2</a></li>
<li style="text-align: justify;"><a href="https://ccb-compute.cs.uni-saarland.de/mirmaster" target="_blank">miRMaster</a></li>
</ul>
</div>
<p> </p>
<p> </p>',
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'info3' => '<p>The counting or expression level calculation is the last step of the processing to generate an expression level matrix. We recommend using MGcount*, a counting tool developed at Diagenode with a suitable annotations file that include the small RNA transcripts that are object of study. MGcount, built on top of featureCounts, employs a flexible quantification approach to deal with datasets containing multiple RNA biotypes such as D-plex libraries. Non-coding RNAs varying in length, biogenesis, and function, may overlap in a genomic region, and are sometimes present in the genome with a high copy number. Consequently, reads may align equally well to more than one position in the reference genome or/and align to a position where more than one annotated transcript is located. MGcount employs two strategies to quantify these reads. Firstly, it assigns reads in rounds prioritizing small-RNAs over long-RNAs ensuring the unbiased read attribution to intergenic and intragenic small RNAs while preventing host-genes transcription over-estimation. Secondly, MGcount collapses loci where reads consistently multi-map into communities of loci with a graph-based approach. The resultant communities are groups of biologically related loci with nearly identical sequence. Subsequently, quantification of reads is performed at the loci-community level, reducing multi-alignments ambiguity at individual locus. Ultimately, this strategy maximizes the transcriptome information analysed and improves the interrogation of non-coding RNAs. MGcount software repository provides annotation files for A. thaliana, H. sapiens, M. musculus and C. elegans. The required input files for MGcount are a .txt file listing the paths to the alignment input files (.bam format) and the annotations file (.gtf format). The output directory path has to be provided as an input as well.</p>
<p><strong>Example command:</strong></p>
<p style="background-color: #f0f0f0;">MGcount -T 2 --gtf Homo_sapiens.GRCh38.gtf --outdir outputs --bam_infiles input_bamfilenames.txt</p>
<p>MGcount provides the choice to enable/disable the quantification of all RNA biotypes included in the annotation file in the form of "communities" as optional parameters for small-RNA (--ml_flag_small) and long-RNA (--ml_flag_long). Both are enabled by default. The main output of MGcount is the count_matrix.csv file containing an expression matrix that can be imported to R or any other software for downstream analyses.<br />A full user guide for MGCount is available here: <a href="https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html">https://filedn.com/lTnUWxFTA93JTyX3Hvbdn2h/mgcount/UserGuide.html</a>.</p>
<p>Other standard counting tools such as featureCounts or HTSeq-counts can also be used alternatively. Given the high complexity of D-Plex libraries, we recommend having a clear understanding of the scientific question and the goal of the project before proceeding to the choice of the counting method as this will strongly impact downstream analyses. Diagenode provides bioinformatics support as a service (please, contact us if you need help with data analysis).</p>
<p>Notice that D-Plex produces forward-stranded data. Stranded libraries have the benefit that reads map to the genome strand where they were originated from. Therefore, when estimating transcript expression, reads aligned to the forward strand should be assigned only to transcript features in the forward strand whereas reads aligned to the reverse strand should be assigned only to transcript features in the reverse strand. For this, make sure you select “stranded mode” in any tool of choice. Stranded mode is selected by default in MGcount.</p>
<p><em><small>*Hita, A., Brocart, G., Fernandez, A. et al. MGcount: a total RNA-seq quantification tool to address multi-mapping and multi-overlapping alignments ambiguity in non-coding transcripts. BMC Bioinformatics 23, 39 (2022). <a href="https://doi.org/10.1186/s12859-021-04544-3">https://doi.org/10.1186/s12859-021-04544-3</a></small></em></p>',
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'meta_description' => 'Small RNA-seq library preparation for Illumina sequencing - Unique D-Plex technology - Optimized for ultra-low input (100 pg total RNA) - UMI reduce PCR biases - UDI detect index hopping - Compatibility with plasma samples - User-friendly and fast protocol',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set A includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li>C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-B" title="D-Plex 24 Single Indexes for Illumina - Set #B">C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C" title="D-Plex 24 Single Indexes for Illumina - Set #C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D" title="D-Plex 24 Single Indexes for Illumina - Set #D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<p style="text-align: left;"><span>D-Plex Unique Dual Indexes Module - Set B includes primer pairs with 24 unique dual barcodes (unique i5 and i7 indexes) for library multiplexing with the <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24" target="_blank" title="D-Plex Small RNA-seq Kit">D-Plex Small RNA-seq Kit</a>. </span></p>
<p style="text-align: left;"><span>The use of UDI is highly recommended to mitigate errors introduced by read misassignment, including index hopping frequently observed with patterned flow cells such as Illumina’s NovaSeq system.</span></p>
<p><span>Four sets are available separately: </span></p>
<ul>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-A">C05030021 - D-Plex Unique Dual Indexes for Illumina - Set A</a></li>
<li>C05030022 - D-Plex Unique Dual Indexes for Illumina - Set B</li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-C">C05030023 - D-Plex Unique Dual Indexes for Illumina - Set C</a></li>
<li><a href="https://www.diagenode.com/en/p/D-Plex-24-Unique-Dual-Indexes-Set-D">C05030024 - D-Plex Unique Dual Indexes for Illumina - Set D</a></li>
</ul>
<p><span>Each set can be used for library multiplexing up to 24. <span>Set A, B, C and D can be used simultaneously for library multiplexing up to 96.</span></span></p>
<p><span>Read more about the </span><a href="https://www.diagenode.com/en/pages/dplex" target="_blank">D-Plex technology</a><span>.</span><span> </span></p>',
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<li>Compatible with <a href="https://www.diagenode.com/en/p/D-Plex-Small-RNA-seq-Library-Prep-x24">D-Plex Small RNA-seq kit for Illumina</a> </li>
<li>Multiplexing up to <strong>96 samples</strong> when combining Set A, B, C and D</li>
<li>Allows for identification of index hopping</li>
</ul>',
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'info2' => '<p><span>D-Plex RNA-seq UDI library constructs bear the TruSeq (Illumina) HT adapters. In case of a multiplexing scenario, it is therefore recommended to submit the D-Plex libraries as TruSeq HT libraries to your sequencing provider. </span><span>Further details are provided in the D-Plex Unique Dual Indexes Module<span> </span><a href="https://www.diagenode.com/files/products/kits/dplex-unique-dual-indexes-manual.pdf" target="_blank">manual</a>.</span></p>',
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'name' => '次世代シーケンシング',
'description' => '<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2 style="font-size: 22px;">DNA断片化、ライブラリー調製、自動化：NGSのワンストップショップ</h2>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-12 medium-12 large-12 columns">
<h4>1. 断片化装置を選択してください：150 bp〜75 kbの範囲でDNAを断片化します。</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-pico-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/bioruptor_pico.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/megaruptor2-1-unit"><img src="https://www.diagenode.com/img/product/shearing_technologies/B06010001_megaruptor2.jpg" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/bioruptor-one-sonication-device"><img src="https://www.diagenode.com/img/product/shearing_technologies/br-one-profil.png" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns">5μlまで断片化：150 bp〜2 kb<br />NGS DNAライブラリー調製およびFFPE核酸抽出に最適で、</td>
<td class="small-4 medium-4 large-4 columns">2 kb〜75 kbの範囲をできます。<br />メイトペアライブラリー調製および長いフラグメントDNAシーケンシングに最適で、この軽量デスクトップデバイスで</td>
<td class="small-4 medium-4 large-4 columns">20または50μlの断片化が可能です。</td>
</tr>
</tbody>
</table>
<table class="small-12 medium-12 large-12 columns">
<tbody>
<tr>
<th class="small-8 medium-8 large-8 columns">
<h4>2. 最適化されたライブラリー調整キットを選択してください。</h4>
</th>
<th class="small-4 medium-4 large-4 columns">
<h4>3. ライブラリー前処理自動化を選択して、比類のないデータ再現性を実感</h4>
</th>
</tr>
<tr style="background-color: #ffffff;">
<td class="small-12 medium-12 large-12 columns"></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns"><a href="../p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><img src="https://www.diagenode.com/img/product/kits/microPlex_library_preparation.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/ideal-library-preparation-kit-x24-incl-index-primer-set-1-24-rxns"><img src="https://www.diagenode.com/img/product/kits/box_kit.jpg" style="display: block; margin-left: auto; margin-right: auto;" height="173" width="250" /></a></td>
<td class="small-4 medium-4 large-4 columns"><a href="../p/sx-8g-ip-star-compact-automated-system-1-unit"><img src="https://www.diagenode.com/img/product/automation/B03000002%20_ipstar_compact.png" style="display: block; margin-left: auto; margin-right: auto;" /></a></td>
</tr>
<tr>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">50pgの低入力：MicroPlex Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">5ng以上：iDeal Library Preparation Kit</td>
<td class="small-4 medium-4 large-4 columns" style="text-align: center;">Achieve great NGS data easily</td>
</tr>
</tbody>
</table>
</div>
</div>
<blockquote>
<div class="row">
<div class="small-12 medium-12 large-12 columns"><span class="label" style="margin-bottom: 16px; margin-left: -22px; font-size: 15px;">DiagenodeがNGS研究にぴったりなプロバイダーである理由</span>
<p>Diagenodeは15年以上もエピジェネティクス研究に専念、専門としています。 ChIP研究クロマチン用のユニークな断片化システムの開発から始まり、 専門知識を活かし、5μlのせん断体積まで可能で、NGS DNAライブラリーの調製に最適な最先端DNA断片化装置の開発にたどり着きました。 我々は以来、ChIP-seq、Methyl-seq、NGSライブラリー調製用キットを研究開発し、業界をリードする免疫沈降研究と同様に、ライブラリー調製を自動化および完結させる独自の自動化システムを開発にも成功しました。</p>
<ul>
<li>信頼されるせん断装置</li>
<li>様々なインプットからのライブラリ作成キット</li>
<li>独自の自動化デバイス</li>
</ul>
</div>
</div>
</blockquote>
<div class="row">
<div class="small-12 columns">
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#panel1a">次世代シーケンシングへの理解とその専門知識</a>
<div id="panel1a" class="content">
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<p><strong>次世代シーケンシング (NGS)</strong> ）は、著しいスケールとハイスループットでシーケンシングを行い、1日に数十億もの塩基生成を可能にします。 NGSのハイスループットは迅速でありながら正確で、再現性のあるデータセットを実現し、さらにシーケンシング費用を削減します。 NGSは、ゲノムシーケンシング、ゲノム再シーケンシング、デノボシーケンシング、トランスクリプトームシーケンシング、その他にDNA-タンパク質相互作用の検出やエピゲノムなどを示します。 指数関数的に増加するシーケンシングデータの需要は、計算分析の障害や解釈、データストレージなどの課題を解決します。</p>
<p>アプリケーションおよび出発物質に応じて、数百万から数十億の鋳型DNA分子を大規模に並行してシーケンシングすることが可能です。その為に、異なる化学物質を使用するいくつかの市販のNGSプラットフォームを利用することができます。 NGSプラットフォームの種類によっては、事前準備とライブラリー作成が必要です。</p>
<p>NGSにとっても、特にデータ処理と分析に関した大きな課題はあります。第3世代技術はゲノミクス研究にさらに革命を起こすであろうと大きく期待されています。</p>
</div>
</div>
<div class="row">
<div class="small-6 medium-6 large-6 columns">
<p><strong>NGS アプリケーション</strong></p>
<ul>
<li>全ゲノム配列決定</li>
<li>デノボシーケンシング</li>
<li>標的配列</li>
<li>Exomeシーケンシング</li>
<li>トランスクリプトーム配列決定</li>
<li>ゲノム配列決定</li>
<li>ミトコンドリア配列決定</li>
<li>DNA-タンパク質相互作用（ChIP-seq</li>
<li>バリアント検出</li>
<li>ゲノム仕上げ</li>
</ul>
</div>
<div class="small-6 medium-6 large-6 columns">
<p><strong>研究分野におけるNGS:</strong></p>
<ul>
<li>腫瘍学</li>
<li>リプロダクティブ・ヘルス</li>
<li>法医学ゲノミクス</li>
<li>アグリゲノミックス</li>
<li>複雑な病気</li>
<li>微生物ゲノミクス</li>
<li>食品・環境ゲノミクス</li>
<li>創薬ゲノミクス - パーソナライズド・メディカル</li>
</ul>
</div>
<div class="small-12 medium-12 large-12 columns">
<p><strong>NGSの用語</strong></p>
<dl>
<dt>リード（読み取り）</dt>
<dd>この装置から得られた連続した単一のストレッチ</dd>
<dt>断片リード</dt>
<dd>フラグメントライブラリからの読み込み。 シーケンシングプラットフォームに応じて、読み取りは通常約100〜300bp。</dd>
<dt>断片ペアエンドリード</dt>
<dd>断片ライブラリーからDNA断片の各末端2つの読み取り。</dd>
<dt>メイトペアリード</dt>
<dd>大きなDNA断片（通常は予め定義されたサイズ範囲）の各末端から2つの読み取り。</dd>
<dt>カバレッジ（例）</dt>
<dd>30×適用範囲とは、参照ゲノム中の各塩基対が平均30回の読み取りを示す。</dd>
</dl>
</div>
</div>
<div class="row">
<div class="small-12 medium-12 large-12 columns">
<h2>NGSプラットフォーム</h2>
<h3><a href="http://www.illumina.com" target="_blank">イルミナ</a></h3>
<p>イルミナは、クローン的に増幅された鋳型DNA（クラスター）上に位置する、蛍光標識された可逆的鎖ターミネーターヌクレオチドを用いた配列別合成技術を使用。 DNAクラスターは、ガラスフローセルの表面上に固定化され、 ワークフローは、4つのヌクレオチド（それぞれ異なる蛍光色素で標識された）の組み込み、4色イメージング、色素や末端基の切断、取り込み、イメージングなどを繰り返します。フローセルは大規模な並列配列決定を受ける。 この方法により、単一蛍光標識されたヌクレオチドの制御添加によるモノヌクレオチドのエラーを回避する可能性があります。 読み取りの長さは、通常約100〜150 bpです。</p>
<h3><a href="http://www.lifetechnologies.com" target="_blank">イオン トレント</a></h3>
<p>イオントレントは、半導体技術チップを用いて、合成中にヌクレオチドを取り込む際に放出されたプロトンを検出します。 これは、イオン球粒子と呼ばれるビーズの表面にエマルションPCR（emPCR）を使用し、リンクされた特定のアダプターを用いてDNA断片を増幅します。 各ビーズは1種類のDNA断片で覆われていて、異なるDNA断片を有するビーズは次いで、チップの陽子感知ウェル内に配置されます。 チップには一度に4つのヌクレオチドのうちの1つが浸水し、このプロセスは異なるヌクレオチドで15秒ごとに繰り返されます。 配列決定の間に4つの塩基の各々が1つずつ導入されます、組み込みの場合はプロトンが放出され、電圧信号が取り込みに比例して検出されます。.</p>
<h3><a href="http://www.pacificbiosciences.com" target="_blank">パシフィック バイオサイエンス</a></h3>
<p>パシフィックバイオサイエンスでは、20kbを超える塩基対の読み取りも、単一分子リアルタイム（SMRT）シーケンシングによる構造および細胞タイプの変化を観察することができます。 このプラットフォームでは、超長鎖二本鎖DNA（dsDNA）断片が、Megaruptor（登録商標）のようなDiagenode装置を用いたランダムシアリングまたは目的の標的領域の増幅によって生成されます。 SMRTbellライブラリーは、ユニバーサルヘアピンアダプターをDNA断片の各末端に連結することによって生成します。 サイズ選択条件による洗浄ステップの後、配列決定プライマーをSMRTbellテンプレートにアニーリングし、鋳型DNAに結合したDNAポリメラーゼを含む配列決定を、蛍光標識ヌクレオチドの存在下で開始。 各塩基が取り込まれると、異なる蛍光のパルスをリアルタイムで検出します。</p>
<h3><a href="https://nanoporetech.com" target="_blank">オックスフォード ナノポア</a></h3>
<p>Oxford Nanoporeは、単一のDNA分子配列決定に基づく技術を開発します。その技術により生物学的分子、すなわちDNAが一群の電気抵抗性高分子膜として位置するナノスケールの孔（ナノ細孔）またはその近くを通過し、イオン電流が変化します。 この変化に関する情報は、例えば4つのヌクレオチド（AまたはG r CまたはT）ならびに修飾されたヌクレオチドすべてを区別することによって分子情報に訳されます。 シーケンシングミニオンデバイスのフローセルは、数百個のナノポアチャネルのセンサアレイを含みます。 DNAサンプルは、Diagenode社のMegaruptor（登録商標）を用いてランダムシアリングによって生成され得る超長鎖DNAフラグメントが必要です。</p>
<h3><a href="http://www.lifetechnologies.com/be/en/home/life-science/sequencing/next-generation-sequencing/solid-next-generation-sequencing.html" target="_blank">SOLiD</a></h3>
<p>SOLiDは、ユニークな化学作用により、何千という個々のDNA分子の同時配列決定を可能にします。 それは、アダプター対ライブラリーのフラグメントが適切で、せん断されたゲノムDNAへのアダプターのライゲーションによるライブラリー作製から始まります。 次のステップでは、エマルジョンPCR（emPCR）を実施して、ビーズの表面上の個々の鋳型DNA分子をクローン的に増幅。 emPCRでは、個々の鋳型DNAをPCR試薬と混合し、水中油型エマルジョン内の疎水性シェルで囲まれた水性液滴内のプライマーコートビーズを、配列決定のためにロードするスライドガラスの表面にランダムに付着。 この技術は、シークエンシングプライマーへのライゲーションで競合する4つの蛍光標識されたジ塩基プローブのセットを使用します。</p>
<h3><a href="http://454.com/products/technology.asp" target="_blank">454</a></h3>
<p>454は、大規模並列パイロシーケンシングを利用しています。 始めに全ゲノムDNAまたは標的遺伝子断片の300〜800bp断片のライブラリー調製します。 次に、DNAフラグメントへのアダプターの付着および単一のDNA鎖の分離。 その後アダプターに連結されたDNAフラグメントをエマルジョンベースのクローン増幅（emPCR）で処理し、DNAライブラリーフラグメントをミクロンサイズのビーズ上に配置します。 各DNA結合ビーズを光ファイバーチップ上のウェルに入れ、器具に挿入します。 4つのDNAヌクレオチドは、配列決定操作中に固定された順序で連続して加えられ、並行して配列決定されます。</p>
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<div class="small-12 medium-12 large-12 columns">
<p><span style="font-weight: 400;">Most of the major next-generation sequencing platforms require ligation of specific adaptor oligos to </span><a href="../applications/dna-rna-shearing"><span style="font-weight: 400;">fragmented DNA or RNA</span></a><span style="font-weight: 400;"> prior to sequencing</span></p>
<p><span style="font-weight: 400;">After input DNA has been fragmented, it is end-repaired and blunt-ended</span><span style="font-weight: 400;">. The next step is a A-tailing in which dAMP is added to the 3´ end of the blunt phosphorylated DNA fragments to prevent concatemerization and to allow the ligation of adaptors with complementary dT overhangs. In addition, barcoded adapters can be incorporated to facilitate multiplexing prior to or during amplification.</span></p>
<center><img src="https://www.diagenode.com/img/categories/library-prep/flux.png" /></center>
<p><span style="font-weight: 400;">Diagenode offers a comprehensive product portfolio for library preparation:<br /></span></p>
<strong><a href="https://www.diagenode.com/en/categories/Library-preparation-for-RNA-seq">D-Plex RNA-seq Library Preparation Kits</a></strong><br />
<p><span style="font-weight: 400;">Diagenode’s new RNA-sequencing solutions utilize the innovative c</span><span style="font-weight: 400;">apture and a</span><span style="font-weight: 400;">mplification by t</span><span style="font-weight: 400;">ailing and s</span><span style="font-weight: 400;">witching”</span><span style="font-weight: 400;">, a ligation-free method to produce DNA libraries for next generation sequencing from low input amounts of RNA. </span><span style="font-weight: 400;"></span><a href="../categories/Library-preparation-for-RNA-seq">Learn more</a></p>
<strong><a href="../categories/library-preparation-for-ChIP-seq">ChIP-seq and DNA sequencing library preparation solutions</a></strong><br />
<p><span style="font-weight: 400;">Our kits have been optimized for DNA library preparation used for next generation sequencing for a wide range of inputs. Using a simple three-step protocols, our</span><a href="http://www.diagenode.com/p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns"><span style="font-weight: 400;"> </span></a><span style="font-weight: 400;">kits are an optimal choice for library preparation from DNA inputs down to 50 pg. </span><a href="../categories/library-preparation-for-ChIP-seq">Learn more</a></p>
<a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span><strong>Bioruptor Pico - short fragments</strong></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">Our well-cited Bioruptor Pico is the shearing device of choice for chromatin and DNA fragmentation. Obtain uniform and tight fragment distributions between 150bp -2kb. </span><a href="../p/bioruptor-pico-sonication-device">Learn more</a></p>
<strong><a href="../p/megaruptor2-1-unit"><span href="../p/bioruptor-pico-sonication-device">Megaruptor</span>® - long fragments</a></strong><a href="../p/bioruptor-pico-sonication-device"><span style="font-weight: 400;"></span></a><a href="../categories/library-preparation-for-ChIP-seq-and-DNA-sequencing"><span style="font-weight: 400;"></span></a><br />
<p><span style="font-weight: 400;"></span><span style="font-weight: 400;">The Megaruptor is designed to shear DNA from 3kb-75kb for long-read sequencing. <a href="../p/megaruptor2-1-unit">Learn more</a></span></p>
<span href="../p/bioruptor-pico-sonication-device"></span><span style="font-weight: 400;"></span></div>
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'description' => '<p>Manganese (Mn) is an essential ubiquitous transition metal and, when occupationally or environmentally overexposed, a well-known risk factor for several neurological pathologies. However, the molecular mechanisms underlying Mn-induced neurotoxicity are largely unknown. In this study, addressing RNA-Seq analysis, bioavailability and survival assays, key pathways of transcriptional responses to Mn overexposure were investigated in the model organism (), providing insights into the Mn-induced cellular stress and damage response. Comparative transcriptome analyses identified a large number of differentially expressed genes (DEGs) in nematodes exposed to MnCl, and functional annotation suggested oxidative nucleotide damage, unfolded protein response and innate immunity as major damage response pathways. Additionally, a time-dependent increase in the transcriptional response after MnCl exposure was identified by means of increased numbers of DEGs, indicating a time-dependent response and activation of the stress responses in Mn neurotoxicity. The data provided here represent a powerful transcriptomic resource in the field of Mn toxicity, and therefore, this study provides a useful basis for further planning of targeted mechanistic studies of Mn-induced neurotoxicity that are urgently needed in the face of increasing industrially caused environmental pollution with Mn.</p>',
'date' => '2022-09-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/36142660',
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'description' => '<p>Interleukin (IL)-33 is a member of the IL-1 cytokine family and is secreted by fibroblasts, endothelial cells, and epithelial cells in tissues (lung, skin, and gastrointestinal) that are exposed to the environment. 1 IL-33 is stored in the nucleus and environmental allergens trigger RIPK1-caspase 8 ripoptosome activation in epithelial cells to enable IL-33 maturation and release. 2 IL-33 signals via its receptor ST2, which is found on multiple immune cells, such as eosinophils, mast cells, T cells, macrophages, basophils, and type 2 innate lymphoid cells, which all promote type-2 innate immune reactions. 3 Therefore, IL-33 plays a central role in promoting allergic inflammatory responses in diseases such as eosinophilic esophagitis (EoE), allergy to food/inhalants, asthma, and atopic dermatitis. Genome-wide association data of these conditions consistently report polymorphisms in the IL-33 gene as a risk factor. 4 Thus, IL-33 has been proposed as a therapeutic target for these conditions. In animal models, IL-33 has been mechanistically linked with allergic, autoimmune, rheumatologic, and inflammatory bowel diseases. 5 However, monogenic human diseases caused by either gain-of-function or loss-of-function variants in the IL-33 gene have not been reported previously.</p>',
'date' => '2022-04-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/35489436',
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'description' => '<p>BACKGROUND: MicroRNA (miRNA) play a significant role in the pathogenesis of complex neurodegenerative diseases including age-related macular degeneration (AMD), acting as post-transcriptional gene suppressors through their association with argonaute 2 (AGO2) - a key member of the RNA Induced Silencing Complex (RISC). Identifying the retinal miRNA/mRNA interactions in health and disease will provide important insight into the key pathways miRNA regulate in disease pathogenesis and may lead to potential therapeutic targets to mediate retinal degeneration. METHODS: To identify the active miRnome targetome interactions in the healthy and degenerating retina, AGO2 HITS-CLIP was performed using a rodent model of photoreceptor degeneration. Analysis of publicly available single-cell RNA sequencing (scRNAseq) data was performed to identify the cellular location of AGO2 and key members of the microRNA targetome in the retina. AGO2 findings were verified by in situ hybridization (RNA) and immunohistochemistry (protein). RESULTS: Analysis revealed a similar miRnome between healthy and damaged retinas, however, a shift in the active targetome was observed with an enrichment of miRNA involvement in inflammatory pathways. This shift was further demonstrated by a change in the seed binding regions of miR-124-3p, the most abundant retinal AGO2-bound miRNA, and has known roles in regulating retinal inflammation. Additionally, photoreceptor cluster miR-183/96/182 were all among the most highly abundant miRNA bound to AGO2. Following damage, AGO2 expression was localized to the inner retinal layers and more in the OLM than in healthy retinas, indicating a locational miRNA response to retinal damage. CONCLUSIONS: This study provides important insight into the alteration of miRNA regulatory activity that occurs as a response to retinal degeneration and explores the miRNA-mRNA targetome as a consequence of retinal degenerations. Further characterisation of these miRNA/mRNA interactions in the context of the degenerating retina may provide an important insight into the active role these miRNA may play in diseases such as AMD.</p>',
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'description' => '<div class="row"><div class="small-12 medium-4 large-4 columns"><div class="panel"><h3><em>lncRNAs in cancer</em></h3><p class="text-left">Gastrointestinal cancer: Gastrointestinal cancers occur as a result of dysregulated signaling pathways and cellular processes, such as the cell cycle or apoptosis. LncRNAs can regulate proliferation of gastrointestinal cancer through interacting with RNA targets, localization to chromatin, or binding to proteins. Read more about Long non-coding RNAs: crucial regulators of gastrointestinal cancer cell proliferation</p><h3><em>microRNAs in cancer</em></h3><p class="text-left">miR-155 has been found overexpressed in many cancer types including hematopoietic cancers, breast, lung and colon cancer<br /><br /> <a href="https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf">https://www.cell.com/trends/molecular-medicine/pdf/S1471-4914(14)00101-4.pdf</a></p></div></div><div class="small-12 medium-8 large-8 columns"><center><img src="https://www.diagenode.com/img/cancer/long-non-coding-rna.jpg" width="550" height="345" /></center><p></p><p>Various non-coding RNAs (ncRNAs) have been found to function as key regulators for transcription, chromatin remodelling, and post-transcriptional modification, thus making them relevant in oncogenesis, both as tumor suppressors and as drivers. For example, a number of microRNAs (miRNAs), one of the more well-studied types of ncRNAs, have been identified as potential cancer biomarkers and clinical targets. Other ncRNAs, such as long noncoding RNAs are involved in cancer regulation and proliferation. Understanding the role of ncRNAs will be critical for cancer research and therapeutic development.</p></div></div>',
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'name' => 'Functional microRNA targetome undergoes degeneration-induced shift inthe retina.',
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'description' => '<p>BACKGROUND: MicroRNA (miRNA) play a significant role in the pathogenesis of complex neurodegenerative diseases including age-related macular degeneration (AMD), acting as post-transcriptional gene suppressors through their association with argonaute 2 (AGO2) - a key member of the RNA Induced Silencing Complex (RISC). Identifying the retinal miRNA/mRNA interactions in health and disease will provide important insight into the key pathways miRNA regulate in disease pathogenesis and may lead to potential therapeutic targets to mediate retinal degeneration. METHODS: To identify the active miRnome targetome interactions in the healthy and degenerating retina, AGO2 HITS-CLIP was performed using a rodent model of photoreceptor degeneration. Analysis of publicly available single-cell RNA sequencing (scRNAseq) data was performed to identify the cellular location of AGO2 and key members of the microRNA targetome in the retina. AGO2 findings were verified by in situ hybridization (RNA) and immunohistochemistry (protein). RESULTS: Analysis revealed a similar miRnome between healthy and damaged retinas, however, a shift in the active targetome was observed with an enrichment of miRNA involvement in inflammatory pathways. This shift was further demonstrated by a change in the seed binding regions of miR-124-3p, the most abundant retinal AGO2-bound miRNA, and has known roles in regulating retinal inflammation. Additionally, photoreceptor cluster miR-183/96/182 were all among the most highly abundant miRNA bound to AGO2. Following damage, AGO2 expression was localized to the inner retinal layers and more in the OLM than in healthy retinas, indicating a locational miRNA response to retinal damage. CONCLUSIONS: This study provides important insight into the alteration of miRNA regulatory activity that occurs as a response to retinal degeneration and explores the miRNA-mRNA targetome as a consequence of retinal degenerations. Further characterisation of these miRNA/mRNA interactions in the context of the degenerating retina may provide an important insight into the active role these miRNA may play in diseases such as AMD.</p>',
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'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/34465369',
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