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<li>Including control DNA and primers to <span>monitor the efficiency of the assay</span>
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'description' => '<p>Diagenode’s<span> </span><b>IPure</b><b><span> </span>kit<span> </span></b>is the only DNA purification kit using magnetic beads, that is specifically optimized for extracting DNA from<span> </span><b>ChIP</b><b>,<span> </span></b><b>MeDIP</b><span> </span>and<span> </span><b>CUT&Tag</b>. The use of the magnetic beads allows for a clear separation of DNA and increases therefore the reproducibility of your DNA purification. This simple and straightforward protocol delivers pure DNA ready for any downstream application (e.g. next generation sequencing). Comparing to phenol-chloroform extraction, the IPure technology has the advantage of being nontoxic and much easier to be carried out on multiple samples.</p>
<center>
<h4>High DNA recovery after purification of ChIP samples using IPure technology</h4>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-chromatin-function.png" width="500" /></center>
<p></p>
<p><small>ChIP assays were performed using different amounts of U2OS cells and the H3K9me3 antibody (Cat. No.<span> </span><span>C15410056</span>; 2 g/IP). <span>The purified DNA was eluted in 50 µl of water and quantified with a Nanodrop.</span></small></p>
<p></p>
<p><strong>Benefits of the IPure kit:</strong></p>
<ul>
<li style="text-align: left;">Provides pure DNA for any downstream application (e. g. Next generation sequencing)</li>
<li style="text-align: left;">Non-toxic</li>
<li style="text-align: left;">Fast & easy to use</li>
<li style="text-align: left;">Optimized for DNA purification after ChIP, MeDIP and CUT&Tag</li>
<li style="text-align: left;">Compatible with automation</li>
<li style="text-align: left;">Validated on the IP-Star Compact</li>
</ul>
</center>',
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'info1' => '<h2>IPure after ChIP</h2>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-A.png" alt="ChIP-seq figure A" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-B.png" alt="ChIP-seq figure B" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-C.png" alt="ChIP-seq figure C" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><small><strong>Figure 1.</strong> Chromatin Immunoprecipitation has been performed using chromatin from HeLa cells, the iDeal ChIP-seq kit for Transcription Factors (containing the IPure module for DNA purification) and the Diagenode ChIP-seq-grade HDAC1 (A), LSD1 (B) and p53 antibody (C). The IP'd DNA was subsequently analysed on an Illumina® Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. This figure shows the peak distribution in regions of chromosome 3 (A), chromosome 12 (B) and chromosome 6 (C) respectively.</small></p>
<p></p>
<h2>IPure after CUT&Tag</h2>
<p>Successful CUT&Tag results showing a low background with high region-specific enrichment has been generated using 50.000 of K562 cells, 1 µg of H3K4me3 or H3K27me3 antibody (Diagenode, C15410003 or C15410069, respectively) and proteinA-Tn5 (1:250) (Diagenode, C01070001). 1 µg of IgG (C15410206) was used as negative control. Samples were purified using the IPure kit v2 or phenol-chloroform purification. The below figures present the comparison of two purification methods.</p>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-fig2.png" style="display: block; margin-left: auto; margin-right: auto;" width="400" /></center><center>
<p style="text-align: center;"><small><strong>Figure 2.</strong> Heatmap 3kb upstream and downstream of the TSS for H3K4me3</small></p>
</center>
<p></p>
<p><img src="https://www.diagenode.com/img/product/kits/ipure-fig3.png" style="display: block; margin-left: auto; margin-right: auto;" width="600" /></p>
<p></p>
<center><small><strong>Figure 3.</strong> Integrative genomics viewer (IGV) visualization of CUT&Tag experiments using Diagenode’s pA-Tn5 transposase (Cat. No. C01070002), H3K27me3 antibody (Cat. No. C15410069) and IPure kit v2 vs phenol chloroform purification (PC).</small></center>
<p></p>
<p></p>
<h2>IPure after MeDIP</h2>
<center><img src="https://www.diagenode.com/img/product/kits/magmedip-seq-figure_multi3.jpg" alt="medip sequencing coverage" width="600" /></center><center></center><center>
<p></p>
<small><strong>Figure 4.</strong> Consistent coverage and methylation detection from different starting amounts of DNA with the Diagenode MagMeDIP-seq Package (including the Ipure kit for DNA purification). Samples containing decreasing starting amounts of DNA (from the top down: 1000 ng (red), 250 ng (blue), 100 ng (green)) originating from human blood were prepared, revealing a consistent coverage profile for the three different starting amounts, which enables reproducible methylation detection. The CpG islands (CGIs) (marked by yellow boxes in the bottom track) are predominantly unmethylated in the human genome, and as expected, we see a depletion of reads at and around CGIs.</small></center>
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'label2' => 'iPure Workflow',
'info2' => '<h2 style="text-align: center;">Kit Method Overview & Time table</h2>
<p><img src="https://www.diagenode.com/img/product/kits/workflow-ipure-cuttag.png" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<h3><strong>Workflow description</strong></h3>
<h5><strong>IPure after ChIP</strong></h5>
<p><strong>Step 1:</strong> Chromatin is decrosslinked and eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added.<br /> <strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet.<br /> <strong>Step 3:</strong> Proteins and remaining buffer are washed away.<br /> <strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after MeDIP</strong></h5>
<p><strong>Step 1:</strong> DNA is eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Remaining buffer are washed away.<br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after CUT&Tag</strong></h5>
<p><strong>Step 1:</strong> pA-Tn5 is inactivated and DNA released from the cells. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Proteins and remaining buffer are washed away. <br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).</p>
<p></p>
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'name' => '5-hydroxymethylcytosine (5-hmC) Antibody (mouse) ',
'description' => '<p>One of the <strong>only two monoclonal antibodies raised against 5-hydroxymethylcytosine (5-hmC).</strong> 5-hmC is a recently discovered DNA modification which results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. Preliminary results indicate that 5-hmC may have important roles distinct from 5-methylcytosine (5-mC). Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics.</p>
<p><strong></strong></p>',
'label1' => 'Validation Data',
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<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig1.png" alt="ChIP" width="160" caption="false" height="280" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 1. An hydroxymethylated DNA IP (hMeDIP) was performed using the Diagenode mouse monoclonal antibody directed against 5-hydroxymethylcytosine (Cat. No. MAb-31HMC-020, MAb-31HMC-050, MAb-31HMC-100).</strong> <br />The IgG isotype antibodies from mouse (Cat. No. kch-819-015) was used as negative control. The DNA was prepared with the GenDNA module of the hMeDIP kit and sonicated with our Bioruptor® (UCD-200/300 series) to have DNA fragments of 300-500 bp. 1 μg of human Hela cells DNA were spiked with non-methylated, methylated, and hydroxymethylated PCR fragments. The IP’d material has been analysed by qPCR using the primer pair specific for the 3 different control sequences. The obtained results show that the mouse monoclonal for 5-hmC is highly specific for this base modification (no IP with non-methylated or methylated C bases containing fragments). </small></p>
</div>
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<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig2.png" alt="ELISA" width="190" caption="false" style="display: block; margin-left: auto; margin-right: auto;" /></p>
</div>
<div class="small-8 columns">
<p><small><strong> Figure 2. Determination of the 5-hmC mouse monoclonal antibody titer </strong><br />To determine the titer, an ELISA was performed using a serial dilution of the Diagenode mouse monoclonal antibody directed against 5-hmC (Cat No. MAb-31HMC-050, MAb-31HMC-100) in antigen coated wells. The antigen used was KHL coupled to 5-hmC base. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:40,000. </small></p>
</div>
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<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig3.png" alt="Dot Blot" width="100" caption="false" height="137" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 3. Dotblot analysis of the Diagenode 5-hmC mouse monoclonal antibody with the C, mC and hmC PCR controls </strong><br />200 to 2 ng (equivalent of 10 to 0.1 pmol of C-bases) of the hmC (1), mC (2) and C (3) PCR controls from the Diagenode “5-hmC, 5-mC & cytosine DNA Standard Pack” (Cat No. AF-101-0020) were spotted on a membrane (Amersham Hybond-N+). The membrane was incubated with 2 μg/ml of the mouse 5-hydroxymethylcytosine monoclonal antibody (dilution 1:500). The membranes were exposed for 30 seconds. </small></p>
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'info2' => '<p>5-hydroxymethylcytosine (5-hmC) has been recently discovered in mammalian DNA. This results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. So far, the 5-hmC bases have been identified in Purkinje neurons, in granule cells and embryonic stem cells where they are present at high levels (up to 0,6% of total nucleotides in Purkinje cells).</p>
<p>Preliminary results indicate that 5-hmC may have important roles distinct from 5-mC. Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics : 5-hydroxymethylcytosine may well represent a new pathway to demethylate DNA involving a repair mechanism converting 5-hmC to cytosine and, as such open up entirely new perspectives in epigenetic studies.</p>
<p>Due to the structural similarity between 5-mC and 5-hmC, these bases are experimentally almost indistinguishable. Recent articles demonstrated that the most common approaches (e.g. enzymatic approaches, bisulfite sequencing) do not account for 5-hmC. The development of the affinity-based technologies appears to be the most powerful way to differentially and specifically enrich 5-mC and 5-hmC sequences. The results shown here illustrate the use of this unique monoclonal antibody against 5-hydroxymethylcytosine that has been fully validated in various technologies.</p>',
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<p><em><strong>CAUTION:</strong> The hydroxymethylated spike-in control is produced from a genomic sequence from Arabidopsis thaliana and may therefore interfere with plant samples. However, it does not show significant homology with other samples species (e.g. human, mouse or rat).</em></p>',
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<div class="small-12 medium-3 large-3 columns"><center><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank"><img src="https://www.diagenode.com/img/banners/banner-nature-publication-580.png" /></a></center></div>
<div class="small-12 medium-9 large-9 columns">
<h3>Sensitive tumour detection and classification using plasma cell-free DNA methylomes<br /><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank">Read the publication</a></h3>
<h3 class="c-article-title u-h1" data-test="article-title" itemprop="name headline">Preparation of cfMeDIP-seq libraries for methylome profiling of plasma cell-free DNA<br /><a href="https://www.nature.com/articles/s41596-019-0202-2" target="_blank" title="cfMeDIP-seq Nature Method">Read the method</a></h3>
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<div class="row">
<div class="large-12 columns"><span>The Methylated DNA Immunoprecipitation is based on the affinity purification of methylated and hydroxymethylated DNA using, respectively, an antibody directed against 5-methylcytosine (5-mC) in the case of MeDIP or 5-hydroxymethylcytosine (5-hmC) in the case of hMeDIP.</span><br />
<h2></h2>
<h2>How it works</h2>
<p>In brief, Methyl DNA IP is performed as follows: Genomic DNA from cultured cells or tissues is prepared, sheared, and then denatured. Then, immunoselection and immunoprecipitation can take place using the antibody directed against 5 methylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d methylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or next generation sequencing.</p>
<h2>Applications</h2>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-kit-x48-48-rxns" class="center alert radius button"> qPCR analysis</a></div>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-seq-package-V2-x10" class="center alert radius button"> NGS analysis </a></div>
<h2>Advantages</h2>
<ul style="font-size: 19px;" class="nobullet">
<li><i class="fa fa-arrow-circle-right"></i> <strong>Unaffected</strong> DNA</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>High enrichment</strong> yield</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>Robust</strong> & <strong>reproducible</strong> techniques</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>NGS</strong> compatible</li>
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<h2></h2>
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<h2>Complete solutions for DNA methylation studies</h2>
<p>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements.</p>
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<p>DNA methylation was the first discovered epigenetic mark and is the most widely studied topic in epigenetics. <em>In vivo</em>, DNA is methylated following DNA replication and is involved in a number of biological processes including the regulation of imprinted genes, X chromosome inactivation. and tumor suppressor gene silencing in cancer cells. Methylation often occurs in cytosine-guanine rich regions of DNA (CpG islands), which are commonly upstream of promoter regions.</p>
</div>
<div class="small-12 medium-12 large-12 columns"><br /><br />
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#dnamethyl"><i class="fa fa-caret-right"></i> Learn more</a>
<div id="dnamethyl" class="content">5-methylcytosine (5-mC) has been known for a long time as the only modification of DNA for epigenetic regulation. In 2009, however, Kriaucionis discovered a second methylated cytosine, 5-hydroxymethylcytosine (5-hmC). The so-called 6th base, is generated by enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine by the TET family of oxygenases. Early reports suggested that 5-hmC may represent an intermediate of active demethylation in a new pathway which demethylates DNA, converting 5-mC to cytosine. Recent evidence fuel this hypothesis suggesting that further oxidation of the hydroxymethyl group leads to a formyl or carboxyl group followed by either deformylation or decarboxylation. The formyl and carboxyl groups of 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC) could be enzymatically removed without excision of the base.
<p class="text-center"><img src="https://www.diagenode.com/img/categories/kits_dna/dna_methylation_variants.jpg" /></p>
</div>
</li>
</ul>
<br />
<h2>Main DNA methylation technologies</h2>
<p style="text-align: justify;">Overview of the <span style="font-weight: 400;">three main approaches for studying DNA methylation.</span></p>
<div class="row">
<ol>
<li style="font-weight: 400;"><span style="font-weight: 400;">Chemical modification with bisulfite – Bisulfite conversion</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Enrichment of methylated DNA (including MeDIP and MBD)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Treatment with methylation-sensitive or dependent restriction enzymes</span></li>
</ol>
<p><span style="font-weight: 400;"> </span></p>
<div class="row">
<table>
<thead>
<tr>
<th></th>
<th>Description</th>
<th width="350">Features</th>
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<td><strong>Bisulfite conversion</strong></td>
<td><span style="font-weight: 400;">Chemical conversion of unmethylated cytosine to uracil. Methylated cytosines are protected from this conversion allowing to determine DNA methylation at single nucleotide resolution.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Single nucleotide resolution</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Quantitative analysis - methylation rate (%)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Gold standard and well studied</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><b>Methylated DNA enrichment</b></td>
<td><span style="font-weight: 400;">(Hydroxy-)Methylated DNA is enriched by using specific antibodies (hMeDIP or MeDIP) or proteins (MBD) that specifically bind methylated CpG sites in fragmented genomic DNA.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Resolution depends on the fragment size of the enriched methylated DNA (300 bp)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Qualitative analysis</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><strong>Restriction enzyme-based digestion</strong></td>
<td><span style="font-weight: 400;">Use of (hydroxy)methylation-sensitive or (hydroxy)methylation-dependent restriction enzymes for DNA methylation analysis at specific sites.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Determination of methylation status is limited by the enzyme recognition site</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Easy to use</span></li>
</ul>
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<div class="row">
<div class="large-12 columns">The hydroxymethylated DNA IP (hMeDIP) is based on the affinity purification of methylated DNA using an antibody directed against 5-hydroxymethylcytosine (5-hmC).
<h3>How it works</h3>
In brief, hydroxymethyl DNA IP is performed as follows: starting from sheared genomic DNA from cultured cells or tissues, the immunoselection and immunoprecipitation can take place using the antibody directed against 5-hydroxymethylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d hydroxymethylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or Next Generation Sequencing.
<h3>Overview</h3>
<p class="text-center"><img src="https://www.diagenode.com/img/applications/magnetic_medip_overview.jpg" caption="false" width="726" height="916" /></p>
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<p><span>The Diagenode hMeDIP kit is designed to immunoprecipitate hydroxymethylated DNA (hMethyl DNA IP). This kit is the first and only example of MeDIP kit specifically designed and fully validated for affinity-capture and detection of hydroxymethylated regions using the highly specific rat, mouse or rabbit antibodies against 5-hmC. </span></p>
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'description' => '<p>Cytosine hydroxymethylation was recently discovered as an important epigenetic mechanism. This cytosine base modification results from the enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) by the TET family of oxygenases. Though the precise role of 5-hmC is the subject of intense research and debate, early studies strongly indicate that it is also involved in gene regulation and in numerous important biological processes including embryonic development, cellular differentiation, stem cell reprogramming and carcinogenesis.</p>
<p>The study of 5-hmC has long been limited due to the lack of high quality, validated tools and technologies that discriminate hydroxymethylation from methylation in regulating gene expression. The use of highly specific antibodies against 5-hmC for the immunoprecipitation of hydroxymethylated DNA offers a reliable solution for hydroxymethylation profiling.</p>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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'name' => 'Developmental and Injury-induced Changes in DNA Methylation inRegenerative versus Non-regenerative Regions of the VertebrateCentral Nervous System',
'authors' => 'Reverdatto S. et al.',
'description' => '<p>Background Because some of its CNS neurons (e.g., retinal ganglion cells after optic nerve crush (ONC)) regenerate axons throughout life, whereas others (e.g., hindbrain neurons after spinal cord injury (SCI)) lose this capacity as tadpoles metamorphose into frogs, the South African claw-toed frog, Xenopus laevis, offers unique opportunities for exploring differences between regenerative and non-regenerative responses to CNS injury within the same organism. An earlier, three-way RNA-seq study (frog ONC eye, tadpole SCI hindbrain, frog SCI hindbrain) identified genes that regulate chromatin accessibility among those that were differentially expressed in regenerative vs non-regenerative CNS [11]. The current study used whole genome bisulfite sequencing (WGBS) of DNA collected from these same animals at the peak period of axon regeneration to study the extent to which DNA methylation could potentially underlie differences in chromatin accessibility between regenerative and non-regenerative CNS. Results Consistent with the hypothesis that DNA of regenerative CNS is more accessible than that of non-regenerative CNS, DNA from both the regenerative tadpole hindbrain and frog eye was less methylated than that of the non-regenerative frog hindbrain. Also, consistent with observations of CNS injury in mammals, DNA methylation in non-regenerative frog hindbrain decreased after SCI. However, contrary to expectations that the level of DNA methylation would decrease even further with axotomy in regenerative CNS, DNA methylation in these regions instead increased with injury. Injury-induced differences in CpG methylation in regenerative CNS became especially enriched in gene promoter regions, whereas non-CpG methylation differences were more evenly distributed across promoter regions, intergenic, and intragenic regions. In non-regenerative CNS, tissue-related (i.e., regenerative vs. non-regenerative CNS) and injury-induced decreases in promoter region CpG methylation were significantly correlated with increased RNA expression, but the injury-induced, increased CpG methylation seen in regenerative CNS across promoter regions was not, suggesting it was associated with increased rather than decreased chromatin accessibility. This hypothesis received support from observations that in regenerative CNS, many genes exhibiting increased, injury-induced, promoter-associated CpG-methylation also exhibited increased RNA expression and association with histone markers for active promoters and enhancers. DNA immunoprecipitation for 5hmC in optic nerve regeneration found that the promoter-associated increases seen in CpG methylation were distinct from those exhibiting changes in 5hmC. Conclusions Although seemingly paradoxical, the increased injury-associated DNA methylation seen in regenerative CNS has many parallels in stem cells and cancer. Thus, these axotomy-induced changes in DNA methylation in regenerative CNS provide evidence for a novel epigenetic state favoring successful over unsuccessful CNS axon regeneration. The datasets described in this study should help lay the foundations for future studies of the molecular and cellular mechanisms involved. The insights gained should, in turn, help point the way to novel therapeutic approaches for treating CNS injury in mammals. Supplementary Information The online version contains supplementary material available at 10.1186/s12864-021-08247-0.</p>',
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'description' => '<p>Aging is the predominant risk factor for atherosclerosis, the leading cause of death. Rare smooth muscle cells (SMCs) clonally expand giving rise to up to ∼70\% of atherosclerotic plaque cells; however, the effect of age on SMC clonality is not known. Our results indicate that aging induces SMC polyclonality and worsens atherosclerosis through non-cell autonomous effects of aged bone marrow-derived cells. Indeed, in myeloid cells from aged mice and humans, TET2 levels are reduced which epigenetically silences integrin β3 resulting in increased cytokine (e.g., tumor necrosis factor [TNF]-α) signaling. In turn, TNFα induces recruitment and expansion of multiple SMCs into the atherosclerotic plaque. Recent studies demonstrate that normal aging is characterized by somatic mutations and clonal expansion of epithelial cells of diverse tissues (e.g., esophagus, endometrium, skin); extrapolating beyond atherogenesis, our results call for future studies evaluating the role of aged myeloid cells in regulating this epithelial cell clonal expansion.</p>',
'date' => '2022-01-01',
'pmid' => 'https://doi.org/10.1101%2F2022.01.18.476756',
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'description' => '<p>Estrogen hormones are implicated in a majority of breast cancers and estrogen receptor alpha (ER), the main nuclear factor mediating estrogen signaling, orchestrates a complex molecular circuitry that is not yet fully elucidated. Here, we investigated genome-wide DNA methylation, histone acetylation and transcription after estradiol (E2) deprivation and re-stimulation to better characterize the ability of ER to coordinate gene regulation. We found that E2 deprivation mostly resulted in DNA hypermethylation and histone deacetylation in enhancers. Transcriptome analysis revealed that E2 deprivation leads to a global down-regulation in gene expression, and more specifically of TET2 demethylase that may be involved in the DNA hypermethylation following short-term E2 deprivation. Further enrichment analysis of transcription factor (TF) binding and motif occurrence highlights the importance of ER connection mainly with two partner TF families, AP-1 and FOX. These interactions take place in the proximity of E2 deprivation-mediated differentially methylated and histone acetylated enhancers. Finally, while most deprivation-dependent epigenetic changes were reversed following E2 re-stimulation, DNA hypermethylation and H3K27 deacetylation at certain enhancers were partially retained. Overall, these results show that inactivation of ER mediates rapid and mostly reversible epigenetic changes at enhancers, and bring new insight into early events, which may ultimately lead to endocrine resistance.</p>',
'date' => '2021-09-01',
'pmid' => 'https://doi.org/10.1093%2Fnar%2Fgkab697',
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'name' => 'Transcription of synaptic plasticity-related genes in patients with somnipathy combined with type 2 diabetes',
'authors' => 'Yi Zhang, Rui Ma, Shaohong Zou, Gaiyu Tong, Gulibakeranmu Abula, Manna Hu, Qing Dai',
'description' => '<p>Objective: To investigate DNA methylation and hydroxymethylation in patients with somnipathy combined with type 2 diabetes, and to explore the effects of DNA methylation and hydroxymethylation on gene expression.</p>
<p>Methods: Thirty patients with somnipathy combined with type 2 diabetes and 20 patients with type-2 diabetes but without somnipathy were considered. DNA methylation of Disks Large Homolog 4 (DLG4) and Ras-related protein Rab-11 (Rab11) was detected by bisulfite sequencing and DNA hydroxymethylation of activity-regulated cytoskeleton-associated protein (Arc), Cyclic AMP-Responsive Element-Binding protein 3 (CREB3) and Early Growth Response protein 1 (EGR1) was analyzed by CHIP analysis. Transcription levels of DLG4, Rab11, Arc, CREB3 and EGR1 were detected by quantitative real-time RT-PCR (qRT-PCR).</p>
<p>Results: Methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 in patients with somnipathy were significantly higher than those in control group (p<0.01). Increased transcription levels of DLG4, Arc and Erg1, and decreased transcription levels of Rab11 and Creb3 were found in patients with somnipathy than in patients without somnipathy. Transcription level of DLG4 was positively, and Rab11 was negatively correlated with their methylation levels. Transcription levels of Arc and Erg1 were positively, and transcription level of Creb3 was negatively correlated with hydroxymethylation levels.</p>
<p>Conclusion: Increased methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 were related to the development of type 2 diabetes in patients with somnipathy. Methylation and hydroxymethylation can significantly affect gene expression at transcription level.</p>',
'date' => '2017-09-03',
'pmid' => 'http://webcache.googleusercontent.com/search?q=cache:t-cxqi84UCcJ:www.alliedacademies.org/articles/transcription-of-synaptic-plasticityrelated-genes-in-patients-with-somnipathy-combined-with-type-2-diabetes.pdf+&cd=1&hl=en&ct=clnk&gl=us',
'doi' => '',
'modified' => '2017-09-25 08:51:27',
'created' => '2017-09-25 08:44:51',
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(int) 4 => array(
'id' => '3265',
'name' => 'Emerging Role of One-Carbon Metabolism and DNA Methylation Enrichment on δ-Containing GABAA Receptor Expression in the Cerebellum of Subjects with Alcohol Use Disorders (AUD',
'authors' => 'Gatta E. et al.',
'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
'date' => '2017-08-19',
'pmid' => 'https://academic.oup.com/ijnp/article/doi/10.1093/ijnp/pyx075/4085582/Emerging-role-of-one-carbon-metabolism-and-DNA',
'doi' => '',
'modified' => '2017-10-09 16:11:05',
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'id' => '3251',
'name' => 'Coordinate Regulation of TET2 and EBNA2 Control DNA Methylation State of Latent Epstein-Barr Virus',
'authors' => 'Lu F. et al.',
'description' => '<p>Epstein-Barr Virus (EBV) latency and its associated carcinogenesis are regulated by dynamic changes in DNA methylation of both virus and host genomes. We show here that the Ten-Eleven Translocation 2 (TET2) gene, implicated in hydroxymethylation and active DNA demethylation, is a key regulator of EBV latency type DNA methylation patterning. EBV latency types are defined by DNA methylation patterns that restrict expression of viral latency genes. We show that TET2 mRNA and protein expression correlate with the highly demethylated EBV type III latency program permissive for expression of EBNA2, EBNA3s, and LMP transcripts. We show that shRNA depletion of TET2 results in a decrease in latency gene expression, but can also trigger a switch to lytic gene expression. TET2 depletion results in the loss of hydroxymethylated cytosine, and corresponding increase in cytosine methylation at key regulatory regions on the viral and host genomes. This also corresponded to a loss of RBP-jκ binding, and decreased histone H3K4 trimethylation at these sites. Furthermore, we show that the TET2 gene, itself, is regulated similar to the EBV genome. ChIP-Seq revealed that TET2 gene contains EBNA2-dependent RBP-jκ and EBF1 binding sites, and is subject to DNA methylation associated transcriptional silencing similar to EBV latency type III genomes. Finally, we provide evidence that TET2 colocalizes with EBNA2-EBF1-RBP-jκ binding sites, and can interact with EBNA2 by co-immunoprecipitation. Taken together, these findings indicate that TET2 gene transcripts are regulated similarly to EBV type III latency genes, and that TET2 protein is a cofactor of EBNA2 and co-regulator of the EBV type III latency program and DNA methylation state..<b>IMPORTANCE</b> Epstein-Barr Virus (EBV) latency and carcinogenesis involves the selective epigenetic modification of viral and cellular genes. Here, we show that TET2, a cellular tumor suppressor involved in active DNA demethylation, plays a central role in regulating DNA methylation state during EBV latency. TET2 is coordinately regulated and functionally interacts with the viral oncogene EBNA2. TET2 and EBNA2 function cooperatively to demethylate genes important for EBV-driven B cells growth transformation.</p>',
'date' => '2017-08-07',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28794029',
'doi' => '',
'modified' => '2017-09-26 09:54:39',
'created' => '2017-09-26 09:54:39',
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(int) 6 => array(
'id' => '3172',
'name' => 'Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer',
'authors' => 'Vafadar-Isfahani N. et al.',
'description' => '<p>Hypomethylation of LINE-1 repeats in cancer has been proposed as the main mechanism behind their activation; this assumption, however, was based on findings from early studies that were biased toward young and transpositionally active elements. Here, we investigate the relationship between methylation of 2 intergenic, transpositionally inactive LINE-1 elements and expression of the LINE-1 chimeric transcript (LCT) 13 and LCT14 driven by their antisense promoters (L1-ASP). Our data from DNA modification, expression, and 5'RACE analyses suggest that colorectal cancer methylation in the regions analyzed is not always associated with LCT repression. Consistent with this, in HCT116 colorectal cancer cells lacking DNA methyltransferases DNMT1 or DNMT3B, LCT13 expression decreases, while cells lacking both DNMTs or treated with the DNMT inhibitor 5-azacytidine (5-aza) show no change in LCT13 expression. Interestingly, levels of the H4K20me3 histone modification are inversely associated with LCT13 and LCT14 expression. Moreover, at these LINE-1s, H4K20me3 levels rather than DNA methylation seem to be good predictor of their sensitivity to 5-aza treatment. Therefore, by studying individual LINE-1 promoters we have shown that in some cases these promoters can be active without losing methylation; in addition, we provide evidence that other factors (e.g., H4K20me3 levels) play prominent roles in their regulation.</p>',
'date' => '2017-03-16',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28300471',
'doi' => '',
'modified' => '2017-05-10 16:26:24',
'created' => '2017-05-10 16:26:24',
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(int) 7 => array(
'id' => '2951',
'name' => 'Maternal immune activation induces GAD1 and GAD2 promoter remodeling in the offspring prefrontal cortex',
'authors' => 'Labouesse MA et al.',
'description' => '<p>Maternal infection during pregnancy increases the risk of neurodevelopmental disorders in the offspring. In addition to its influence on other neuronal systems, this early-life environmental adversity has been shown to negatively affect cortical γ-aminobutyric acid (GABA) functions in adult life, including impaired prefrontal expression of enzymes required for GABA synthesis. The underlying molecular processes, however, remain largely unknown. In the present study, we explored whether epigenetic modifications represent a mechanism whereby maternal infection during pregnancy can induce such GABAergic impairments in the offspring. We used an established mouse model of prenatal immune challenge that is based on maternal treatment with the viral mimetic poly(I:C). We found that prenatal immune activation increased prefrontal levels of 5-methylated cytosines (5mC) and 5-hydroxymethylated cytosines (5hmC) in the promoter region of GAD1, which encodes the 67-kDa isoform of the GABA-synthesising enzyme glutamic acid decarboxylase (GAD67). The early-life challenge also increased 5mC levels at the promoter region of GAD2, which encodes the 65-kDa GAD isoform (GAD65). These effects were accompanied by elevated GAD1 and GAD2 promoter binding of methyl CpG-binding protein 2 (MeCP2) and by reduced GAD67 and GAD65 mRNA expression. Moreover, the epigenetic modifications at the GAD1 promoter correlated with prenatal infection-induced impairments in working memory and social interaction. Our study thus highlights that hypermethylation of GAD1 and GAD2 promoters may be an important molecular mechanism linking prenatal infection to presynaptic GABAergic impairments and associated behavioral and cognitive abnormalities in the offspring.</p>',
'date' => '2015-12-02',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26575259',
'doi' => ' 10.1080/15592294.2015.1114202',
'modified' => '2016-06-10 16:32:32',
'created' => '2016-06-10 16:32:32',
'ProductsPublication' => array(
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(int) 8 => array(
'id' => '2950',
'name' => 'Hepatic DNA hydroxymethylation is site-specifically altered by chronic alcohol consumption and aging',
'authors' => 'Tammen SA et al.',
'description' => '<div class="">
<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
</div>',
'date' => '2015-11-14',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26578530',
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<a href="/cn/p/dna-hydroxymethylation-control-package-48-rxns"><img src="/img/product/kits/methyl-kit-icon.png" alt="Methylation kit icon" class="th"/></a> </div>
<div class="small-12 columns">
<div class="small-6 columns" style="padding-left:0px;padding-right:0px;margin-top:-6px;margin-left:-1px">
<span class="success label" style="">C02040018</span>
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<p>将 <input name="data[Cart][quantity]" placeholder="1" value="1" min="1" style="width:60px;display:inline" type="number" id="CartQuantity" required="required"/> <strong> DNA hydroxymethylation control package</strong> 添加至我的购物车。</p>
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<h6 style="height:60px">DNA hydroxymethylation control package</h6>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
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<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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<center>
<h4>High DNA recovery after purification of ChIP samples using IPure technology</h4>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-chromatin-function.png" width="500" /></center>
<p></p>
<p><small>ChIP assays were performed using different amounts of U2OS cells and the H3K9me3 antibody (Cat. No.<span> </span><span>C15410056</span>; 2 g/IP). <span>The purified DNA was eluted in 50 µl of water and quantified with a Nanodrop.</span></small></p>
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<p><strong>Benefits of the IPure kit:</strong></p>
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<li style="text-align: left;">Non-toxic</li>
<li style="text-align: left;">Fast & easy to use</li>
<li style="text-align: left;">Optimized for DNA purification after ChIP, MeDIP and CUT&Tag</li>
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<li style="text-align: left;">Validated on the IP-Star Compact</li>
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<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-A.png" alt="ChIP-seq figure A" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-B.png" alt="ChIP-seq figure B" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-C.png" alt="ChIP-seq figure C" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><small><strong>Figure 1.</strong> Chromatin Immunoprecipitation has been performed using chromatin from HeLa cells, the iDeal ChIP-seq kit for Transcription Factors (containing the IPure module for DNA purification) and the Diagenode ChIP-seq-grade HDAC1 (A), LSD1 (B) and p53 antibody (C). The IP'd DNA was subsequently analysed on an Illumina® Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. This figure shows the peak distribution in regions of chromosome 3 (A), chromosome 12 (B) and chromosome 6 (C) respectively.</small></p>
<p></p>
<h2>IPure after CUT&Tag</h2>
<p>Successful CUT&Tag results showing a low background with high region-specific enrichment has been generated using 50.000 of K562 cells, 1 µg of H3K4me3 or H3K27me3 antibody (Diagenode, C15410003 or C15410069, respectively) and proteinA-Tn5 (1:250) (Diagenode, C01070001). 1 µg of IgG (C15410206) was used as negative control. Samples were purified using the IPure kit v2 or phenol-chloroform purification. The below figures present the comparison of two purification methods.</p>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-fig2.png" style="display: block; margin-left: auto; margin-right: auto;" width="400" /></center><center>
<p style="text-align: center;"><small><strong>Figure 2.</strong> Heatmap 3kb upstream and downstream of the TSS for H3K4me3</small></p>
</center>
<p></p>
<p><img src="https://www.diagenode.com/img/product/kits/ipure-fig3.png" style="display: block; margin-left: auto; margin-right: auto;" width="600" /></p>
<p></p>
<center><small><strong>Figure 3.</strong> Integrative genomics viewer (IGV) visualization of CUT&Tag experiments using Diagenode’s pA-Tn5 transposase (Cat. No. C01070002), H3K27me3 antibody (Cat. No. C15410069) and IPure kit v2 vs phenol chloroform purification (PC).</small></center>
<p></p>
<p></p>
<h2>IPure after MeDIP</h2>
<center><img src="https://www.diagenode.com/img/product/kits/magmedip-seq-figure_multi3.jpg" alt="medip sequencing coverage" width="600" /></center><center></center><center>
<p></p>
<small><strong>Figure 4.</strong> Consistent coverage and methylation detection from different starting amounts of DNA with the Diagenode MagMeDIP-seq Package (including the Ipure kit for DNA purification). Samples containing decreasing starting amounts of DNA (from the top down: 1000 ng (red), 250 ng (blue), 100 ng (green)) originating from human blood were prepared, revealing a consistent coverage profile for the three different starting amounts, which enables reproducible methylation detection. The CpG islands (CGIs) (marked by yellow boxes in the bottom track) are predominantly unmethylated in the human genome, and as expected, we see a depletion of reads at and around CGIs.</small></center>
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<p><img src="https://www.diagenode.com/img/product/kits/workflow-ipure-cuttag.png" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<h3><strong>Workflow description</strong></h3>
<h5><strong>IPure after ChIP</strong></h5>
<p><strong>Step 1:</strong> Chromatin is decrosslinked and eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added.<br /> <strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet.<br /> <strong>Step 3:</strong> Proteins and remaining buffer are washed away.<br /> <strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after MeDIP</strong></h5>
<p><strong>Step 1:</strong> DNA is eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Remaining buffer are washed away.<br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after CUT&Tag</strong></h5>
<p><strong>Step 1:</strong> pA-Tn5 is inactivated and DNA released from the cells. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Proteins and remaining buffer are washed away. <br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).</p>
<p></p>
<p></p>
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'meta_description' => 'IPure kit v2',
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'antibody_id' => '47',
'name' => '5-hydroxymethylcytosine (5-hmC) Antibody (mouse) ',
'description' => '<p>One of the <strong>only two monoclonal antibodies raised against 5-hydroxymethylcytosine (5-hmC).</strong> 5-hmC is a recently discovered DNA modification which results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. Preliminary results indicate that 5-hmC may have important roles distinct from 5-methylcytosine (5-mC). Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics.</p>
<p><strong></strong></p>',
'label1' => 'Validation Data',
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<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig1.png" alt="ChIP" width="160" caption="false" height="280" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 1. An hydroxymethylated DNA IP (hMeDIP) was performed using the Diagenode mouse monoclonal antibody directed against 5-hydroxymethylcytosine (Cat. No. MAb-31HMC-020, MAb-31HMC-050, MAb-31HMC-100).</strong> <br />The IgG isotype antibodies from mouse (Cat. No. kch-819-015) was used as negative control. The DNA was prepared with the GenDNA module of the hMeDIP kit and sonicated with our Bioruptor® (UCD-200/300 series) to have DNA fragments of 300-500 bp. 1 μg of human Hela cells DNA were spiked with non-methylated, methylated, and hydroxymethylated PCR fragments. The IP’d material has been analysed by qPCR using the primer pair specific for the 3 different control sequences. The obtained results show that the mouse monoclonal for 5-hmC is highly specific for this base modification (no IP with non-methylated or methylated C bases containing fragments). </small></p>
</div>
</div>
<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig2.png" alt="ELISA" width="190" caption="false" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 2. Determination of the 5-hmC mouse monoclonal antibody titer </strong><br />To determine the titer, an ELISA was performed using a serial dilution of the Diagenode mouse monoclonal antibody directed against 5-hmC (Cat No. MAb-31HMC-050, MAb-31HMC-100) in antigen coated wells. The antigen used was KHL coupled to 5-hmC base. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:40,000. </small></p>
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</div>
<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig3.png" alt="Dot Blot" width="100" caption="false" height="137" style="display: block; margin-left: auto; margin-right: auto;" /></p>
</div>
<div class="small-8 columns">
<p><small><strong> Figure 3. Dotblot analysis of the Diagenode 5-hmC mouse monoclonal antibody with the C, mC and hmC PCR controls </strong><br />200 to 2 ng (equivalent of 10 to 0.1 pmol of C-bases) of the hmC (1), mC (2) and C (3) PCR controls from the Diagenode “5-hmC, 5-mC & cytosine DNA Standard Pack” (Cat No. AF-101-0020) were spotted on a membrane (Amersham Hybond-N+). The membrane was incubated with 2 μg/ml of the mouse 5-hydroxymethylcytosine monoclonal antibody (dilution 1:500). The membranes were exposed for 30 seconds. </small></p>
</div>
</div>',
'label2' => 'Target description',
'info2' => '<p>5-hydroxymethylcytosine (5-hmC) has been recently discovered in mammalian DNA. This results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. So far, the 5-hmC bases have been identified in Purkinje neurons, in granule cells and embryonic stem cells where they are present at high levels (up to 0,6% of total nucleotides in Purkinje cells).</p>
<p>Preliminary results indicate that 5-hmC may have important roles distinct from 5-mC. Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics : 5-hydroxymethylcytosine may well represent a new pathway to demethylate DNA involving a repair mechanism converting 5-hmC to cytosine and, as such open up entirely new perspectives in epigenetic studies.</p>
<p>Due to the structural similarity between 5-mC and 5-hmC, these bases are experimentally almost indistinguishable. Recent articles demonstrated that the most common approaches (e.g. enzymatic approaches, bisulfite sequencing) do not account for 5-hmC. The development of the affinity-based technologies appears to be the most powerful way to differentially and specifically enrich 5-mC and 5-hmC sequences. The results shown here illustrate the use of this unique monoclonal antibody against 5-hydroxymethylcytosine that has been fully validated in various technologies.</p>',
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'format' => '50 µg/50 µl',
'catalog_number' => 'C15200200-50',
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'meta_title' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) | Diagenode',
'meta_keywords' => '5-hydroxymethylcytosine,monoclonal antibody ,Diagenode',
'meta_description' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) validated in hMeDIP, DB and ELISA. Batch-specific data available on the website. Sample size available.',
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'created' => '2015-06-29 14:08:20',
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(int) 3 => array(
'id' => '3154',
'antibody_id' => null,
'name' => 'DNA hydroxymethylation control package',
'description' => '<p>The DNA hydroxymethylation control package includes one hydroxymethylated spike-in control and its corresponding qPCR primer set that can be added to the DNA sample of interest for any hydroxymethylation profiling experiment (e.g. with Diagenode's <a href="https://www.diagenode.com/en/p/hmedip-kit-x16-monoclonal-mouse-antibody-16-rxns">hMeDIP Kit</a>).</p>
<p><em><strong>CAUTION:</strong> The hydroxymethylated spike-in control is produced from a genomic sequence from Arabidopsis thaliana and may therefore interfere with plant samples. However, it does not show significant homology with other samples species (e.g. human, mouse or rat).</em></p>',
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'description' => '<div class="row extra-spaced">
<div class="small-12 medium-3 large-3 columns"><center><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank"><img src="https://www.diagenode.com/img/banners/banner-nature-publication-580.png" /></a></center></div>
<div class="small-12 medium-9 large-9 columns">
<h3>Sensitive tumour detection and classification using plasma cell-free DNA methylomes<br /><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank">Read the publication</a></h3>
<h3 class="c-article-title u-h1" data-test="article-title" itemprop="name headline">Preparation of cfMeDIP-seq libraries for methylome profiling of plasma cell-free DNA<br /><a href="https://www.nature.com/articles/s41596-019-0202-2" target="_blank" title="cfMeDIP-seq Nature Method">Read the method</a></h3>
</div>
</div>
<div class="row">
<div class="large-12 columns"><span>The Methylated DNA Immunoprecipitation is based on the affinity purification of methylated and hydroxymethylated DNA using, respectively, an antibody directed against 5-methylcytosine (5-mC) in the case of MeDIP or 5-hydroxymethylcytosine (5-hmC) in the case of hMeDIP.</span><br />
<h2></h2>
<h2>How it works</h2>
<p>In brief, Methyl DNA IP is performed as follows: Genomic DNA from cultured cells or tissues is prepared, sheared, and then denatured. Then, immunoselection and immunoprecipitation can take place using the antibody directed against 5 methylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d methylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or next generation sequencing.</p>
<h2>Applications</h2>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-kit-x48-48-rxns" class="center alert radius button"> qPCR analysis</a></div>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-seq-package-V2-x10" class="center alert radius button"> NGS analysis </a></div>
<h2>Advantages</h2>
<ul style="font-size: 19px;" class="nobullet">
<li><i class="fa fa-arrow-circle-right"></i> <strong>Unaffected</strong> DNA</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>High enrichment</strong> yield</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>Robust</strong> & <strong>reproducible</strong> techniques</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>NGS</strong> compatible</li>
</ul>
<h2></h2>
</div>
</div>
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'meta_description' => 'Methylated DNA immunoprecipitation method is based on the affinity purification of methylated DNA using an antibody directed against 5 methylcytosine (5-mC). ',
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<div class="large-12 columns">
<div style="text-align: justify;" class="small-12 medium-8 large-8 columns">
<h2>Complete solutions for DNA methylation studies</h2>
<p>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements.</p>
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<div class="small-12 medium-4 large-4 columns text-center"><a href="../landing-pages/dna-methylation-grant-applications"><img src="https://www.diagenode.com/img/banners/banner-dna-grant.png" alt="" /></a></div>
<div style="text-align: justify;" class="small-12 medium-12 large-12 columns">
<p>DNA methylation was the first discovered epigenetic mark and is the most widely studied topic in epigenetics. <em>In vivo</em>, DNA is methylated following DNA replication and is involved in a number of biological processes including the regulation of imprinted genes, X chromosome inactivation. and tumor suppressor gene silencing in cancer cells. Methylation often occurs in cytosine-guanine rich regions of DNA (CpG islands), which are commonly upstream of promoter regions.</p>
</div>
<div class="small-12 medium-12 large-12 columns"><br /><br />
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#dnamethyl"><i class="fa fa-caret-right"></i> Learn more</a>
<div id="dnamethyl" class="content">5-methylcytosine (5-mC) has been known for a long time as the only modification of DNA for epigenetic regulation. In 2009, however, Kriaucionis discovered a second methylated cytosine, 5-hydroxymethylcytosine (5-hmC). The so-called 6th base, is generated by enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine by the TET family of oxygenases. Early reports suggested that 5-hmC may represent an intermediate of active demethylation in a new pathway which demethylates DNA, converting 5-mC to cytosine. Recent evidence fuel this hypothesis suggesting that further oxidation of the hydroxymethyl group leads to a formyl or carboxyl group followed by either deformylation or decarboxylation. The formyl and carboxyl groups of 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC) could be enzymatically removed without excision of the base.
<p class="text-center"><img src="https://www.diagenode.com/img/categories/kits_dna/dna_methylation_variants.jpg" /></p>
</div>
</li>
</ul>
<br />
<h2>Main DNA methylation technologies</h2>
<p style="text-align: justify;">Overview of the <span style="font-weight: 400;">three main approaches for studying DNA methylation.</span></p>
<div class="row">
<ol>
<li style="font-weight: 400;"><span style="font-weight: 400;">Chemical modification with bisulfite – Bisulfite conversion</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Enrichment of methylated DNA (including MeDIP and MBD)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Treatment with methylation-sensitive or dependent restriction enzymes</span></li>
</ol>
<p><span style="font-weight: 400;"> </span></p>
<div class="row">
<table>
<thead>
<tr>
<th></th>
<th>Description</th>
<th width="350">Features</th>
</tr>
</thead>
<tbody>
<tr>
<td><strong>Bisulfite conversion</strong></td>
<td><span style="font-weight: 400;">Chemical conversion of unmethylated cytosine to uracil. Methylated cytosines are protected from this conversion allowing to determine DNA methylation at single nucleotide resolution.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Single nucleotide resolution</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Quantitative analysis - methylation rate (%)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Gold standard and well studied</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><b>Methylated DNA enrichment</b></td>
<td><span style="font-weight: 400;">(Hydroxy-)Methylated DNA is enriched by using specific antibodies (hMeDIP or MeDIP) or proteins (MBD) that specifically bind methylated CpG sites in fragmented genomic DNA.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Resolution depends on the fragment size of the enriched methylated DNA (300 bp)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Qualitative analysis</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><strong>Restriction enzyme-based digestion</strong></td>
<td><span style="font-weight: 400;">Use of (hydroxy)methylation-sensitive or (hydroxy)methylation-dependent restriction enzymes for DNA methylation analysis at specific sites.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Determination of methylation status is limited by the enzyme recognition site</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Easy to use</span></li>
</ul>
</td>
</tr>
</tbody>
</table>
</div>
</div>
<div class="row"></div>
</div>
</div>
<div class="large-12 columns"></div>
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'meta_description' => 'Complete, optimized solutions for analyzing DNA methylation manually or on our automated system.',
'meta_title' => 'DNA Methylation - Bisulfite sequencing - Epigenetics | Diagenode',
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'name' => 'Hydroxymethylated DNA immunoprecipitation',
'description' => '<p></p>
<div class="row">
<div class="large-12 columns">The hydroxymethylated DNA IP (hMeDIP) is based on the affinity purification of methylated DNA using an antibody directed against 5-hydroxymethylcytosine (5-hmC).
<h3>How it works</h3>
In brief, hydroxymethyl DNA IP is performed as follows: starting from sheared genomic DNA from cultured cells or tissues, the immunoselection and immunoprecipitation can take place using the antibody directed against 5-hydroxymethylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d hydroxymethylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or Next Generation Sequencing.
<h3>Overview</h3>
<p class="text-center"><img src="https://www.diagenode.com/img/applications/magnetic_medip_overview.jpg" caption="false" width="726" height="916" /></p>
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<p><span>The Diagenode hMeDIP kit is designed to immunoprecipitate hydroxymethylated DNA (hMethyl DNA IP). This kit is the first and only example of MeDIP kit specifically designed and fully validated for affinity-capture and detection of hydroxymethylated regions using the highly specific rat, mouse or rabbit antibodies against 5-hmC. </span></p>
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'description' => '<p>Cytosine hydroxymethylation was recently discovered as an important epigenetic mechanism. This cytosine base modification results from the enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) by the TET family of oxygenases. Though the precise role of 5-hmC is the subject of intense research and debate, early studies strongly indicate that it is also involved in gene regulation and in numerous important biological processes including embryonic development, cellular differentiation, stem cell reprogramming and carcinogenesis.</p>
<p>The study of 5-hmC has long been limited due to the lack of high quality, validated tools and technologies that discriminate hydroxymethylation from methylation in regulating gene expression. The use of highly specific antibodies against 5-hmC for the immunoprecipitation of hydroxymethylated DNA offers a reliable solution for hydroxymethylation profiling.</p>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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'name' => 'Developmental and Injury-induced Changes in DNA Methylation inRegenerative versus Non-regenerative Regions of the VertebrateCentral Nervous System',
'authors' => 'Reverdatto S. et al.',
'description' => '<p>Background Because some of its CNS neurons (e.g., retinal ganglion cells after optic nerve crush (ONC)) regenerate axons throughout life, whereas others (e.g., hindbrain neurons after spinal cord injury (SCI)) lose this capacity as tadpoles metamorphose into frogs, the South African claw-toed frog, Xenopus laevis, offers unique opportunities for exploring differences between regenerative and non-regenerative responses to CNS injury within the same organism. An earlier, three-way RNA-seq study (frog ONC eye, tadpole SCI hindbrain, frog SCI hindbrain) identified genes that regulate chromatin accessibility among those that were differentially expressed in regenerative vs non-regenerative CNS [11]. The current study used whole genome bisulfite sequencing (WGBS) of DNA collected from these same animals at the peak period of axon regeneration to study the extent to which DNA methylation could potentially underlie differences in chromatin accessibility between regenerative and non-regenerative CNS. Results Consistent with the hypothesis that DNA of regenerative CNS is more accessible than that of non-regenerative CNS, DNA from both the regenerative tadpole hindbrain and frog eye was less methylated than that of the non-regenerative frog hindbrain. Also, consistent with observations of CNS injury in mammals, DNA methylation in non-regenerative frog hindbrain decreased after SCI. However, contrary to expectations that the level of DNA methylation would decrease even further with axotomy in regenerative CNS, DNA methylation in these regions instead increased with injury. Injury-induced differences in CpG methylation in regenerative CNS became especially enriched in gene promoter regions, whereas non-CpG methylation differences were more evenly distributed across promoter regions, intergenic, and intragenic regions. In non-regenerative CNS, tissue-related (i.e., regenerative vs. non-regenerative CNS) and injury-induced decreases in promoter region CpG methylation were significantly correlated with increased RNA expression, but the injury-induced, increased CpG methylation seen in regenerative CNS across promoter regions was not, suggesting it was associated with increased rather than decreased chromatin accessibility. This hypothesis received support from observations that in regenerative CNS, many genes exhibiting increased, injury-induced, promoter-associated CpG-methylation also exhibited increased RNA expression and association with histone markers for active promoters and enhancers. DNA immunoprecipitation for 5hmC in optic nerve regeneration found that the promoter-associated increases seen in CpG methylation were distinct from those exhibiting changes in 5hmC. Conclusions Although seemingly paradoxical, the increased injury-associated DNA methylation seen in regenerative CNS has many parallels in stem cells and cancer. Thus, these axotomy-induced changes in DNA methylation in regenerative CNS provide evidence for a novel epigenetic state favoring successful over unsuccessful CNS axon regeneration. The datasets described in this study should help lay the foundations for future studies of the molecular and cellular mechanisms involved. The insights gained should, in turn, help point the way to novel therapeutic approaches for treating CNS injury in mammals. Supplementary Information The online version contains supplementary material available at 10.1186/s12864-021-08247-0.</p>',
'date' => '2022-01-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/34979916',
'doi' => '10.1186/s12864-021-08247-0',
'modified' => '2022-05-20 09:20:25',
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'id' => '4271',
'name' => 'Bone marrow age dictates clonality of smooth muscle-derived cells in theatherosclerotic plaque',
'authors' => 'Kabir Inamul et al.',
'description' => '<p>Aging is the predominant risk factor for atherosclerosis, the leading cause of death. Rare smooth muscle cells (SMCs) clonally expand giving rise to up to ∼70\% of atherosclerotic plaque cells; however, the effect of age on SMC clonality is not known. Our results indicate that aging induces SMC polyclonality and worsens atherosclerosis through non-cell autonomous effects of aged bone marrow-derived cells. Indeed, in myeloid cells from aged mice and humans, TET2 levels are reduced which epigenetically silences integrin β3 resulting in increased cytokine (e.g., tumor necrosis factor [TNF]-α) signaling. In turn, TNFα induces recruitment and expansion of multiple SMCs into the atherosclerotic plaque. Recent studies demonstrate that normal aging is characterized by somatic mutations and clonal expansion of epithelial cells of diverse tissues (e.g., esophagus, endometrium, skin); extrapolating beyond atherogenesis, our results call for future studies evaluating the role of aged myeloid cells in regulating this epithelial cell clonal expansion.</p>',
'date' => '2022-01-01',
'pmid' => 'https://doi.org/10.1101%2F2022.01.18.476756',
'doi' => '10.1101/2022.01.18.476756',
'modified' => '2022-05-23 09:45:53',
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'id' => '4329',
'name' => 'Epigenetic remodelling of enhancers in response to estrogen deprivationand re-stimulation.',
'authors' => 'Sklias Athena et al.',
'description' => '<p>Estrogen hormones are implicated in a majority of breast cancers and estrogen receptor alpha (ER), the main nuclear factor mediating estrogen signaling, orchestrates a complex molecular circuitry that is not yet fully elucidated. Here, we investigated genome-wide DNA methylation, histone acetylation and transcription after estradiol (E2) deprivation and re-stimulation to better characterize the ability of ER to coordinate gene regulation. We found that E2 deprivation mostly resulted in DNA hypermethylation and histone deacetylation in enhancers. Transcriptome analysis revealed that E2 deprivation leads to a global down-regulation in gene expression, and more specifically of TET2 demethylase that may be involved in the DNA hypermethylation following short-term E2 deprivation. Further enrichment analysis of transcription factor (TF) binding and motif occurrence highlights the importance of ER connection mainly with two partner TF families, AP-1 and FOX. These interactions take place in the proximity of E2 deprivation-mediated differentially methylated and histone acetylated enhancers. Finally, while most deprivation-dependent epigenetic changes were reversed following E2 re-stimulation, DNA hypermethylation and H3K27 deacetylation at certain enhancers were partially retained. Overall, these results show that inactivation of ER mediates rapid and mostly reversible epigenetic changes at enhancers, and bring new insight into early events, which may ultimately lead to endocrine resistance.</p>',
'date' => '2021-09-01',
'pmid' => 'https://doi.org/10.1093%2Fnar%2Fgkab697',
'doi' => '10.1093/nar/gkab697',
'modified' => '2022-06-22 09:25:09',
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'name' => 'Transcription of synaptic plasticity-related genes in patients with somnipathy combined with type 2 diabetes',
'authors' => 'Yi Zhang, Rui Ma, Shaohong Zou, Gaiyu Tong, Gulibakeranmu Abula, Manna Hu, Qing Dai',
'description' => '<p>Objective: To investigate DNA methylation and hydroxymethylation in patients with somnipathy combined with type 2 diabetes, and to explore the effects of DNA methylation and hydroxymethylation on gene expression.</p>
<p>Methods: Thirty patients with somnipathy combined with type 2 diabetes and 20 patients with type-2 diabetes but without somnipathy were considered. DNA methylation of Disks Large Homolog 4 (DLG4) and Ras-related protein Rab-11 (Rab11) was detected by bisulfite sequencing and DNA hydroxymethylation of activity-regulated cytoskeleton-associated protein (Arc), Cyclic AMP-Responsive Element-Binding protein 3 (CREB3) and Early Growth Response protein 1 (EGR1) was analyzed by CHIP analysis. Transcription levels of DLG4, Rab11, Arc, CREB3 and EGR1 were detected by quantitative real-time RT-PCR (qRT-PCR).</p>
<p>Results: Methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 in patients with somnipathy were significantly higher than those in control group (p<0.01). Increased transcription levels of DLG4, Arc and Erg1, and decreased transcription levels of Rab11 and Creb3 were found in patients with somnipathy than in patients without somnipathy. Transcription level of DLG4 was positively, and Rab11 was negatively correlated with their methylation levels. Transcription levels of Arc and Erg1 were positively, and transcription level of Creb3 was negatively correlated with hydroxymethylation levels.</p>
<p>Conclusion: Increased methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 were related to the development of type 2 diabetes in patients with somnipathy. Methylation and hydroxymethylation can significantly affect gene expression at transcription level.</p>',
'date' => '2017-09-03',
'pmid' => 'http://webcache.googleusercontent.com/search?q=cache:t-cxqi84UCcJ:www.alliedacademies.org/articles/transcription-of-synaptic-plasticityrelated-genes-in-patients-with-somnipathy-combined-with-type-2-diabetes.pdf+&cd=1&hl=en&ct=clnk&gl=us',
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'id' => '3265',
'name' => 'Emerging Role of One-Carbon Metabolism and DNA Methylation Enrichment on δ-Containing GABAA Receptor Expression in the Cerebellum of Subjects with Alcohol Use Disorders (AUD',
'authors' => 'Gatta E. et al.',
'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
'date' => '2017-08-19',
'pmid' => 'https://academic.oup.com/ijnp/article/doi/10.1093/ijnp/pyx075/4085582/Emerging-role-of-one-carbon-metabolism-and-DNA',
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'id' => '3251',
'name' => 'Coordinate Regulation of TET2 and EBNA2 Control DNA Methylation State of Latent Epstein-Barr Virus',
'authors' => 'Lu F. et al.',
'description' => '<p>Epstein-Barr Virus (EBV) latency and its associated carcinogenesis are regulated by dynamic changes in DNA methylation of both virus and host genomes. We show here that the Ten-Eleven Translocation 2 (TET2) gene, implicated in hydroxymethylation and active DNA demethylation, is a key regulator of EBV latency type DNA methylation patterning. EBV latency types are defined by DNA methylation patterns that restrict expression of viral latency genes. We show that TET2 mRNA and protein expression correlate with the highly demethylated EBV type III latency program permissive for expression of EBNA2, EBNA3s, and LMP transcripts. We show that shRNA depletion of TET2 results in a decrease in latency gene expression, but can also trigger a switch to lytic gene expression. TET2 depletion results in the loss of hydroxymethylated cytosine, and corresponding increase in cytosine methylation at key regulatory regions on the viral and host genomes. This also corresponded to a loss of RBP-jκ binding, and decreased histone H3K4 trimethylation at these sites. Furthermore, we show that the TET2 gene, itself, is regulated similar to the EBV genome. ChIP-Seq revealed that TET2 gene contains EBNA2-dependent RBP-jκ and EBF1 binding sites, and is subject to DNA methylation associated transcriptional silencing similar to EBV latency type III genomes. Finally, we provide evidence that TET2 colocalizes with EBNA2-EBF1-RBP-jκ binding sites, and can interact with EBNA2 by co-immunoprecipitation. Taken together, these findings indicate that TET2 gene transcripts are regulated similarly to EBV type III latency genes, and that TET2 protein is a cofactor of EBNA2 and co-regulator of the EBV type III latency program and DNA methylation state..<b>IMPORTANCE</b> Epstein-Barr Virus (EBV) latency and carcinogenesis involves the selective epigenetic modification of viral and cellular genes. Here, we show that TET2, a cellular tumor suppressor involved in active DNA demethylation, plays a central role in regulating DNA methylation state during EBV latency. TET2 is coordinately regulated and functionally interacts with the viral oncogene EBNA2. TET2 and EBNA2 function cooperatively to demethylate genes important for EBV-driven B cells growth transformation.</p>',
'date' => '2017-08-07',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28794029',
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'id' => '3172',
'name' => 'Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer',
'authors' => 'Vafadar-Isfahani N. et al.',
'description' => '<p>Hypomethylation of LINE-1 repeats in cancer has been proposed as the main mechanism behind their activation; this assumption, however, was based on findings from early studies that were biased toward young and transpositionally active elements. Here, we investigate the relationship between methylation of 2 intergenic, transpositionally inactive LINE-1 elements and expression of the LINE-1 chimeric transcript (LCT) 13 and LCT14 driven by their antisense promoters (L1-ASP). Our data from DNA modification, expression, and 5'RACE analyses suggest that colorectal cancer methylation in the regions analyzed is not always associated with LCT repression. Consistent with this, in HCT116 colorectal cancer cells lacking DNA methyltransferases DNMT1 or DNMT3B, LCT13 expression decreases, while cells lacking both DNMTs or treated with the DNMT inhibitor 5-azacytidine (5-aza) show no change in LCT13 expression. Interestingly, levels of the H4K20me3 histone modification are inversely associated with LCT13 and LCT14 expression. Moreover, at these LINE-1s, H4K20me3 levels rather than DNA methylation seem to be good predictor of their sensitivity to 5-aza treatment. Therefore, by studying individual LINE-1 promoters we have shown that in some cases these promoters can be active without losing methylation; in addition, we provide evidence that other factors (e.g., H4K20me3 levels) play prominent roles in their regulation.</p>',
'date' => '2017-03-16',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28300471',
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'name' => 'Maternal immune activation induces GAD1 and GAD2 promoter remodeling in the offspring prefrontal cortex',
'authors' => 'Labouesse MA et al.',
'description' => '<p>Maternal infection during pregnancy increases the risk of neurodevelopmental disorders in the offspring. In addition to its influence on other neuronal systems, this early-life environmental adversity has been shown to negatively affect cortical γ-aminobutyric acid (GABA) functions in adult life, including impaired prefrontal expression of enzymes required for GABA synthesis. The underlying molecular processes, however, remain largely unknown. In the present study, we explored whether epigenetic modifications represent a mechanism whereby maternal infection during pregnancy can induce such GABAergic impairments in the offspring. We used an established mouse model of prenatal immune challenge that is based on maternal treatment with the viral mimetic poly(I:C). We found that prenatal immune activation increased prefrontal levels of 5-methylated cytosines (5mC) and 5-hydroxymethylated cytosines (5hmC) in the promoter region of GAD1, which encodes the 67-kDa isoform of the GABA-synthesising enzyme glutamic acid decarboxylase (GAD67). The early-life challenge also increased 5mC levels at the promoter region of GAD2, which encodes the 65-kDa GAD isoform (GAD65). These effects were accompanied by elevated GAD1 and GAD2 promoter binding of methyl CpG-binding protein 2 (MeCP2) and by reduced GAD67 and GAD65 mRNA expression. Moreover, the epigenetic modifications at the GAD1 promoter correlated with prenatal infection-induced impairments in working memory and social interaction. Our study thus highlights that hypermethylation of GAD1 and GAD2 promoters may be an important molecular mechanism linking prenatal infection to presynaptic GABAergic impairments and associated behavioral and cognitive abnormalities in the offspring.</p>',
'date' => '2015-12-02',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26575259',
'doi' => ' 10.1080/15592294.2015.1114202',
'modified' => '2016-06-10 16:32:32',
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(int) 8 => array(
'id' => '2950',
'name' => 'Hepatic DNA hydroxymethylation is site-specifically altered by chronic alcohol consumption and aging',
'authors' => 'Tammen SA et al.',
'description' => '<div class="">
<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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<p>将 <input name="data[Cart][quantity]" placeholder="1" value="1" min="1" style="width:60px;display:inline" type="number" id="CartQuantity" required="required"/> <strong> IPure kit v2</strong> 添加至我的购物车。</p>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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'authors' => 'Tammen SA et al.',
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<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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<li>Including control DNA and primers to <span>monitor the efficiency of the assay</span>
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<li>hmeDNA and unmethylated DNA sequences and primer pairs</li>
<li>Mouse primer pairs against Sfi1 targeting hydroxymethylated gene in mouse</li>
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</li>
<li>Improved single-tube, magnetic bead-based protocol</li>
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'description' => '<p>Diagenode’s<span> </span><b>IPure</b><b><span> </span>kit<span> </span></b>is the only DNA purification kit using magnetic beads, that is specifically optimized for extracting DNA from<span> </span><b>ChIP</b><b>,<span> </span></b><b>MeDIP</b><span> </span>and<span> </span><b>CUT&Tag</b>. The use of the magnetic beads allows for a clear separation of DNA and increases therefore the reproducibility of your DNA purification. This simple and straightforward protocol delivers pure DNA ready for any downstream application (e.g. next generation sequencing). Comparing to phenol-chloroform extraction, the IPure technology has the advantage of being nontoxic and much easier to be carried out on multiple samples.</p>
<center>
<h4>High DNA recovery after purification of ChIP samples using IPure technology</h4>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-chromatin-function.png" width="500" /></center>
<p></p>
<p><small>ChIP assays were performed using different amounts of U2OS cells and the H3K9me3 antibody (Cat. No.<span> </span><span>C15410056</span>; 2 g/IP). <span>The purified DNA was eluted in 50 µl of water and quantified with a Nanodrop.</span></small></p>
<p></p>
<p><strong>Benefits of the IPure kit:</strong></p>
<ul>
<li style="text-align: left;">Provides pure DNA for any downstream application (e. g. Next generation sequencing)</li>
<li style="text-align: left;">Non-toxic</li>
<li style="text-align: left;">Fast & easy to use</li>
<li style="text-align: left;">Optimized for DNA purification after ChIP, MeDIP and CUT&Tag</li>
<li style="text-align: left;">Compatible with automation</li>
<li style="text-align: left;">Validated on the IP-Star Compact</li>
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<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-A.png" alt="ChIP-seq figure A" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<p><small><strong>Figure 1.</strong> Chromatin Immunoprecipitation has been performed using chromatin from HeLa cells, the iDeal ChIP-seq kit for Transcription Factors (containing the IPure module for DNA purification) and the Diagenode ChIP-seq-grade HDAC1 (A), LSD1 (B) and p53 antibody (C). The IP'd DNA was subsequently analysed on an Illumina® Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. This figure shows the peak distribution in regions of chromosome 3 (A), chromosome 12 (B) and chromosome 6 (C) respectively.</small></p>
<p></p>
<h2>IPure after CUT&Tag</h2>
<p>Successful CUT&Tag results showing a low background with high region-specific enrichment has been generated using 50.000 of K562 cells, 1 µg of H3K4me3 or H3K27me3 antibody (Diagenode, C15410003 or C15410069, respectively) and proteinA-Tn5 (1:250) (Diagenode, C01070001). 1 µg of IgG (C15410206) was used as negative control. Samples were purified using the IPure kit v2 or phenol-chloroform purification. The below figures present the comparison of two purification methods.</p>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-fig2.png" style="display: block; margin-left: auto; margin-right: auto;" width="400" /></center><center>
<p style="text-align: center;"><small><strong>Figure 2.</strong> Heatmap 3kb upstream and downstream of the TSS for H3K4me3</small></p>
</center>
<p></p>
<p><img src="https://www.diagenode.com/img/product/kits/ipure-fig3.png" style="display: block; margin-left: auto; margin-right: auto;" width="600" /></p>
<p></p>
<center><small><strong>Figure 3.</strong> Integrative genomics viewer (IGV) visualization of CUT&Tag experiments using Diagenode’s pA-Tn5 transposase (Cat. No. C01070002), H3K27me3 antibody (Cat. No. C15410069) and IPure kit v2 vs phenol chloroform purification (PC).</small></center>
<p></p>
<p></p>
<h2>IPure after MeDIP</h2>
<center><img src="https://www.diagenode.com/img/product/kits/magmedip-seq-figure_multi3.jpg" alt="medip sequencing coverage" width="600" /></center><center></center><center>
<p></p>
<small><strong>Figure 4.</strong> Consistent coverage and methylation detection from different starting amounts of DNA with the Diagenode MagMeDIP-seq Package (including the Ipure kit for DNA purification). Samples containing decreasing starting amounts of DNA (from the top down: 1000 ng (red), 250 ng (blue), 100 ng (green)) originating from human blood were prepared, revealing a consistent coverage profile for the three different starting amounts, which enables reproducible methylation detection. The CpG islands (CGIs) (marked by yellow boxes in the bottom track) are predominantly unmethylated in the human genome, and as expected, we see a depletion of reads at and around CGIs.</small></center>
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'info2' => '<h2 style="text-align: center;">Kit Method Overview & Time table</h2>
<p><img src="https://www.diagenode.com/img/product/kits/workflow-ipure-cuttag.png" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<h3><strong>Workflow description</strong></h3>
<h5><strong>IPure after ChIP</strong></h5>
<p><strong>Step 1:</strong> Chromatin is decrosslinked and eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added.<br /> <strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet.<br /> <strong>Step 3:</strong> Proteins and remaining buffer are washed away.<br /> <strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after MeDIP</strong></h5>
<p><strong>Step 1:</strong> DNA is eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Remaining buffer are washed away.<br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after CUT&Tag</strong></h5>
<p><strong>Step 1:</strong> pA-Tn5 is inactivated and DNA released from the cells. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Proteins and remaining buffer are washed away. <br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).</p>
<p></p>
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'meta_description' => 'IPure kit v2',
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'name' => '5-hydroxymethylcytosine (5-hmC) Antibody (mouse) ',
'description' => '<p>One of the <strong>only two monoclonal antibodies raised against 5-hydroxymethylcytosine (5-hmC).</strong> 5-hmC is a recently discovered DNA modification which results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. Preliminary results indicate that 5-hmC may have important roles distinct from 5-methylcytosine (5-mC). Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics.</p>
<p><strong></strong></p>',
'label1' => 'Validation Data',
'info1' => '<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig1.png" alt="ChIP" width="160" caption="false" height="280" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 1. An hydroxymethylated DNA IP (hMeDIP) was performed using the Diagenode mouse monoclonal antibody directed against 5-hydroxymethylcytosine (Cat. No. MAb-31HMC-020, MAb-31HMC-050, MAb-31HMC-100).</strong> <br />The IgG isotype antibodies from mouse (Cat. No. kch-819-015) was used as negative control. The DNA was prepared with the GenDNA module of the hMeDIP kit and sonicated with our Bioruptor® (UCD-200/300 series) to have DNA fragments of 300-500 bp. 1 μg of human Hela cells DNA were spiked with non-methylated, methylated, and hydroxymethylated PCR fragments. The IP’d material has been analysed by qPCR using the primer pair specific for the 3 different control sequences. The obtained results show that the mouse monoclonal for 5-hmC is highly specific for this base modification (no IP with non-methylated or methylated C bases containing fragments). </small></p>
</div>
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<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig2.png" alt="ELISA" width="190" caption="false" style="display: block; margin-left: auto; margin-right: auto;" /></p>
</div>
<div class="small-8 columns">
<p><small><strong> Figure 2. Determination of the 5-hmC mouse monoclonal antibody titer </strong><br />To determine the titer, an ELISA was performed using a serial dilution of the Diagenode mouse monoclonal antibody directed against 5-hmC (Cat No. MAb-31HMC-050, MAb-31HMC-100) in antigen coated wells. The antigen used was KHL coupled to 5-hmC base. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:40,000. </small></p>
</div>
</div>
<div class="row">
<div class="small-4 columns">
<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig3.png" alt="Dot Blot" width="100" caption="false" height="137" style="display: block; margin-left: auto; margin-right: auto;" /></p>
</div>
<div class="small-8 columns">
<p><small><strong> Figure 3. Dotblot analysis of the Diagenode 5-hmC mouse monoclonal antibody with the C, mC and hmC PCR controls </strong><br />200 to 2 ng (equivalent of 10 to 0.1 pmol of C-bases) of the hmC (1), mC (2) and C (3) PCR controls from the Diagenode “5-hmC, 5-mC & cytosine DNA Standard Pack” (Cat No. AF-101-0020) were spotted on a membrane (Amersham Hybond-N+). The membrane was incubated with 2 μg/ml of the mouse 5-hydroxymethylcytosine monoclonal antibody (dilution 1:500). The membranes were exposed for 30 seconds. </small></p>
</div>
</div>',
'label2' => 'Target description',
'info2' => '<p>5-hydroxymethylcytosine (5-hmC) has been recently discovered in mammalian DNA. This results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. So far, the 5-hmC bases have been identified in Purkinje neurons, in granule cells and embryonic stem cells where they are present at high levels (up to 0,6% of total nucleotides in Purkinje cells).</p>
<p>Preliminary results indicate that 5-hmC may have important roles distinct from 5-mC. Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics : 5-hydroxymethylcytosine may well represent a new pathway to demethylate DNA involving a repair mechanism converting 5-hmC to cytosine and, as such open up entirely new perspectives in epigenetic studies.</p>
<p>Due to the structural similarity between 5-mC and 5-hmC, these bases are experimentally almost indistinguishable. Recent articles demonstrated that the most common approaches (e.g. enzymatic approaches, bisulfite sequencing) do not account for 5-hmC. The development of the affinity-based technologies appears to be the most powerful way to differentially and specifically enrich 5-mC and 5-hmC sequences. The results shown here illustrate the use of this unique monoclonal antibody against 5-hydroxymethylcytosine that has been fully validated in various technologies.</p>',
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'catalog_number' => 'C15200200-50',
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'meta_title' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) | Diagenode',
'meta_keywords' => '5-hydroxymethylcytosine,monoclonal antibody ,Diagenode',
'meta_description' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) validated in hMeDIP, DB and ELISA. Batch-specific data available on the website. Sample size available.',
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'description' => '<p>The DNA hydroxymethylation control package includes one hydroxymethylated spike-in control and its corresponding qPCR primer set that can be added to the DNA sample of interest for any hydroxymethylation profiling experiment (e.g. with Diagenode's <a href="https://www.diagenode.com/en/p/hmedip-kit-x16-monoclonal-mouse-antibody-16-rxns">hMeDIP Kit</a>).</p>
<p><em><strong>CAUTION:</strong> The hydroxymethylated spike-in control is produced from a genomic sequence from Arabidopsis thaliana and may therefore interfere with plant samples. However, it does not show significant homology with other samples species (e.g. human, mouse or rat).</em></p>',
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<div class="small-12 medium-3 large-3 columns"><center><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank"><img src="https://www.diagenode.com/img/banners/banner-nature-publication-580.png" /></a></center></div>
<div class="small-12 medium-9 large-9 columns">
<h3>Sensitive tumour detection and classification using plasma cell-free DNA methylomes<br /><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank">Read the publication</a></h3>
<h3 class="c-article-title u-h1" data-test="article-title" itemprop="name headline">Preparation of cfMeDIP-seq libraries for methylome profiling of plasma cell-free DNA<br /><a href="https://www.nature.com/articles/s41596-019-0202-2" target="_blank" title="cfMeDIP-seq Nature Method">Read the method</a></h3>
</div>
</div>
<div class="row">
<div class="large-12 columns"><span>The Methylated DNA Immunoprecipitation is based on the affinity purification of methylated and hydroxymethylated DNA using, respectively, an antibody directed against 5-methylcytosine (5-mC) in the case of MeDIP or 5-hydroxymethylcytosine (5-hmC) in the case of hMeDIP.</span><br />
<h2></h2>
<h2>How it works</h2>
<p>In brief, Methyl DNA IP is performed as follows: Genomic DNA from cultured cells or tissues is prepared, sheared, and then denatured. Then, immunoselection and immunoprecipitation can take place using the antibody directed against 5 methylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d methylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or next generation sequencing.</p>
<h2>Applications</h2>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-kit-x48-48-rxns" class="center alert radius button"> qPCR analysis</a></div>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-seq-package-V2-x10" class="center alert radius button"> NGS analysis </a></div>
<h2>Advantages</h2>
<ul style="font-size: 19px;" class="nobullet">
<li><i class="fa fa-arrow-circle-right"></i> <strong>Unaffected</strong> DNA</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>High enrichment</strong> yield</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>Robust</strong> & <strong>reproducible</strong> techniques</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>NGS</strong> compatible</li>
</ul>
<h2></h2>
</div>
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<div class="large-12 columns">
<div style="text-align: justify;" class="small-12 medium-8 large-8 columns">
<h2>Complete solutions for DNA methylation studies</h2>
<p>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements.</p>
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<div class="small-12 medium-4 large-4 columns text-center"><a href="../landing-pages/dna-methylation-grant-applications"><img src="https://www.diagenode.com/img/banners/banner-dna-grant.png" alt="" /></a></div>
<div style="text-align: justify;" class="small-12 medium-12 large-12 columns">
<p>DNA methylation was the first discovered epigenetic mark and is the most widely studied topic in epigenetics. <em>In vivo</em>, DNA is methylated following DNA replication and is involved in a number of biological processes including the regulation of imprinted genes, X chromosome inactivation. and tumor suppressor gene silencing in cancer cells. Methylation often occurs in cytosine-guanine rich regions of DNA (CpG islands), which are commonly upstream of promoter regions.</p>
</div>
<div class="small-12 medium-12 large-12 columns"><br /><br />
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#dnamethyl"><i class="fa fa-caret-right"></i> Learn more</a>
<div id="dnamethyl" class="content">5-methylcytosine (5-mC) has been known for a long time as the only modification of DNA for epigenetic regulation. In 2009, however, Kriaucionis discovered a second methylated cytosine, 5-hydroxymethylcytosine (5-hmC). The so-called 6th base, is generated by enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine by the TET family of oxygenases. Early reports suggested that 5-hmC may represent an intermediate of active demethylation in a new pathway which demethylates DNA, converting 5-mC to cytosine. Recent evidence fuel this hypothesis suggesting that further oxidation of the hydroxymethyl group leads to a formyl or carboxyl group followed by either deformylation or decarboxylation. The formyl and carboxyl groups of 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC) could be enzymatically removed without excision of the base.
<p class="text-center"><img src="https://www.diagenode.com/img/categories/kits_dna/dna_methylation_variants.jpg" /></p>
</div>
</li>
</ul>
<br />
<h2>Main DNA methylation technologies</h2>
<p style="text-align: justify;">Overview of the <span style="font-weight: 400;">three main approaches for studying DNA methylation.</span></p>
<div class="row">
<ol>
<li style="font-weight: 400;"><span style="font-weight: 400;">Chemical modification with bisulfite – Bisulfite conversion</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Enrichment of methylated DNA (including MeDIP and MBD)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Treatment with methylation-sensitive or dependent restriction enzymes</span></li>
</ol>
<p><span style="font-weight: 400;"> </span></p>
<div class="row">
<table>
<thead>
<tr>
<th></th>
<th>Description</th>
<th width="350">Features</th>
</tr>
</thead>
<tbody>
<tr>
<td><strong>Bisulfite conversion</strong></td>
<td><span style="font-weight: 400;">Chemical conversion of unmethylated cytosine to uracil. Methylated cytosines are protected from this conversion allowing to determine DNA methylation at single nucleotide resolution.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Single nucleotide resolution</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Quantitative analysis - methylation rate (%)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Gold standard and well studied</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><b>Methylated DNA enrichment</b></td>
<td><span style="font-weight: 400;">(Hydroxy-)Methylated DNA is enriched by using specific antibodies (hMeDIP or MeDIP) or proteins (MBD) that specifically bind methylated CpG sites in fragmented genomic DNA.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Resolution depends on the fragment size of the enriched methylated DNA (300 bp)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Qualitative analysis</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><strong>Restriction enzyme-based digestion</strong></td>
<td><span style="font-weight: 400;">Use of (hydroxy)methylation-sensitive or (hydroxy)methylation-dependent restriction enzymes for DNA methylation analysis at specific sites.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Determination of methylation status is limited by the enzyme recognition site</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Easy to use</span></li>
</ul>
</td>
</tr>
</tbody>
</table>
</div>
</div>
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</div>
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<div class="row">
<div class="large-12 columns">The hydroxymethylated DNA IP (hMeDIP) is based on the affinity purification of methylated DNA using an antibody directed against 5-hydroxymethylcytosine (5-hmC).
<h3>How it works</h3>
In brief, hydroxymethyl DNA IP is performed as follows: starting from sheared genomic DNA from cultured cells or tissues, the immunoselection and immunoprecipitation can take place using the antibody directed against 5-hydroxymethylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d hydroxymethylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or Next Generation Sequencing.
<h3>Overview</h3>
<p class="text-center"><img src="https://www.diagenode.com/img/applications/magnetic_medip_overview.jpg" caption="false" width="726" height="916" /></p>
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<p><span>The Diagenode hMeDIP kit is designed to immunoprecipitate hydroxymethylated DNA (hMethyl DNA IP). This kit is the first and only example of MeDIP kit specifically designed and fully validated for affinity-capture and detection of hydroxymethylated regions using the highly specific rat, mouse or rabbit antibodies against 5-hmC. </span></p>
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'description' => '<p>Cytosine hydroxymethylation was recently discovered as an important epigenetic mechanism. This cytosine base modification results from the enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) by the TET family of oxygenases. Though the precise role of 5-hmC is the subject of intense research and debate, early studies strongly indicate that it is also involved in gene regulation and in numerous important biological processes including embryonic development, cellular differentiation, stem cell reprogramming and carcinogenesis.</p>
<p>The study of 5-hmC has long been limited due to the lack of high quality, validated tools and technologies that discriminate hydroxymethylation from methylation in regulating gene expression. The use of highly specific antibodies against 5-hmC for the immunoprecipitation of hydroxymethylated DNA offers a reliable solution for hydroxymethylation profiling.</p>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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'name' => 'Developmental and Injury-induced Changes in DNA Methylation inRegenerative versus Non-regenerative Regions of the VertebrateCentral Nervous System',
'authors' => 'Reverdatto S. et al.',
'description' => '<p>Background Because some of its CNS neurons (e.g., retinal ganglion cells after optic nerve crush (ONC)) regenerate axons throughout life, whereas others (e.g., hindbrain neurons after spinal cord injury (SCI)) lose this capacity as tadpoles metamorphose into frogs, the South African claw-toed frog, Xenopus laevis, offers unique opportunities for exploring differences between regenerative and non-regenerative responses to CNS injury within the same organism. An earlier, three-way RNA-seq study (frog ONC eye, tadpole SCI hindbrain, frog SCI hindbrain) identified genes that regulate chromatin accessibility among those that were differentially expressed in regenerative vs non-regenerative CNS [11]. The current study used whole genome bisulfite sequencing (WGBS) of DNA collected from these same animals at the peak period of axon regeneration to study the extent to which DNA methylation could potentially underlie differences in chromatin accessibility between regenerative and non-regenerative CNS. Results Consistent with the hypothesis that DNA of regenerative CNS is more accessible than that of non-regenerative CNS, DNA from both the regenerative tadpole hindbrain and frog eye was less methylated than that of the non-regenerative frog hindbrain. Also, consistent with observations of CNS injury in mammals, DNA methylation in non-regenerative frog hindbrain decreased after SCI. However, contrary to expectations that the level of DNA methylation would decrease even further with axotomy in regenerative CNS, DNA methylation in these regions instead increased with injury. Injury-induced differences in CpG methylation in regenerative CNS became especially enriched in gene promoter regions, whereas non-CpG methylation differences were more evenly distributed across promoter regions, intergenic, and intragenic regions. In non-regenerative CNS, tissue-related (i.e., regenerative vs. non-regenerative CNS) and injury-induced decreases in promoter region CpG methylation were significantly correlated with increased RNA expression, but the injury-induced, increased CpG methylation seen in regenerative CNS across promoter regions was not, suggesting it was associated with increased rather than decreased chromatin accessibility. This hypothesis received support from observations that in regenerative CNS, many genes exhibiting increased, injury-induced, promoter-associated CpG-methylation also exhibited increased RNA expression and association with histone markers for active promoters and enhancers. DNA immunoprecipitation for 5hmC in optic nerve regeneration found that the promoter-associated increases seen in CpG methylation were distinct from those exhibiting changes in 5hmC. Conclusions Although seemingly paradoxical, the increased injury-associated DNA methylation seen in regenerative CNS has many parallels in stem cells and cancer. Thus, these axotomy-induced changes in DNA methylation in regenerative CNS provide evidence for a novel epigenetic state favoring successful over unsuccessful CNS axon regeneration. The datasets described in this study should help lay the foundations for future studies of the molecular and cellular mechanisms involved. The insights gained should, in turn, help point the way to novel therapeutic approaches for treating CNS injury in mammals. Supplementary Information The online version contains supplementary material available at 10.1186/s12864-021-08247-0.</p>',
'date' => '2022-01-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/34979916',
'doi' => '10.1186/s12864-021-08247-0',
'modified' => '2022-05-20 09:20:25',
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'id' => '4271',
'name' => 'Bone marrow age dictates clonality of smooth muscle-derived cells in theatherosclerotic plaque',
'authors' => 'Kabir Inamul et al.',
'description' => '<p>Aging is the predominant risk factor for atherosclerosis, the leading cause of death. Rare smooth muscle cells (SMCs) clonally expand giving rise to up to ∼70\% of atherosclerotic plaque cells; however, the effect of age on SMC clonality is not known. Our results indicate that aging induces SMC polyclonality and worsens atherosclerosis through non-cell autonomous effects of aged bone marrow-derived cells. Indeed, in myeloid cells from aged mice and humans, TET2 levels are reduced which epigenetically silences integrin β3 resulting in increased cytokine (e.g., tumor necrosis factor [TNF]-α) signaling. In turn, TNFα induces recruitment and expansion of multiple SMCs into the atherosclerotic plaque. Recent studies demonstrate that normal aging is characterized by somatic mutations and clonal expansion of epithelial cells of diverse tissues (e.g., esophagus, endometrium, skin); extrapolating beyond atherogenesis, our results call for future studies evaluating the role of aged myeloid cells in regulating this epithelial cell clonal expansion.</p>',
'date' => '2022-01-01',
'pmid' => 'https://doi.org/10.1101%2F2022.01.18.476756',
'doi' => '10.1101/2022.01.18.476756',
'modified' => '2022-05-23 09:45:53',
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'id' => '4329',
'name' => 'Epigenetic remodelling of enhancers in response to estrogen deprivationand re-stimulation.',
'authors' => 'Sklias Athena et al.',
'description' => '<p>Estrogen hormones are implicated in a majority of breast cancers and estrogen receptor alpha (ER), the main nuclear factor mediating estrogen signaling, orchestrates a complex molecular circuitry that is not yet fully elucidated. Here, we investigated genome-wide DNA methylation, histone acetylation and transcription after estradiol (E2) deprivation and re-stimulation to better characterize the ability of ER to coordinate gene regulation. We found that E2 deprivation mostly resulted in DNA hypermethylation and histone deacetylation in enhancers. Transcriptome analysis revealed that E2 deprivation leads to a global down-regulation in gene expression, and more specifically of TET2 demethylase that may be involved in the DNA hypermethylation following short-term E2 deprivation. Further enrichment analysis of transcription factor (TF) binding and motif occurrence highlights the importance of ER connection mainly with two partner TF families, AP-1 and FOX. These interactions take place in the proximity of E2 deprivation-mediated differentially methylated and histone acetylated enhancers. Finally, while most deprivation-dependent epigenetic changes were reversed following E2 re-stimulation, DNA hypermethylation and H3K27 deacetylation at certain enhancers were partially retained. Overall, these results show that inactivation of ER mediates rapid and mostly reversible epigenetic changes at enhancers, and bring new insight into early events, which may ultimately lead to endocrine resistance.</p>',
'date' => '2021-09-01',
'pmid' => 'https://doi.org/10.1093%2Fnar%2Fgkab697',
'doi' => '10.1093/nar/gkab697',
'modified' => '2022-06-22 09:25:09',
'created' => '2022-05-19 10:41:50',
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'id' => '3245',
'name' => 'Transcription of synaptic plasticity-related genes in patients with somnipathy combined with type 2 diabetes',
'authors' => 'Yi Zhang, Rui Ma, Shaohong Zou, Gaiyu Tong, Gulibakeranmu Abula, Manna Hu, Qing Dai',
'description' => '<p>Objective: To investigate DNA methylation and hydroxymethylation in patients with somnipathy combined with type 2 diabetes, and to explore the effects of DNA methylation and hydroxymethylation on gene expression.</p>
<p>Methods: Thirty patients with somnipathy combined with type 2 diabetes and 20 patients with type-2 diabetes but without somnipathy were considered. DNA methylation of Disks Large Homolog 4 (DLG4) and Ras-related protein Rab-11 (Rab11) was detected by bisulfite sequencing and DNA hydroxymethylation of activity-regulated cytoskeleton-associated protein (Arc), Cyclic AMP-Responsive Element-Binding protein 3 (CREB3) and Early Growth Response protein 1 (EGR1) was analyzed by CHIP analysis. Transcription levels of DLG4, Rab11, Arc, CREB3 and EGR1 were detected by quantitative real-time RT-PCR (qRT-PCR).</p>
<p>Results: Methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 in patients with somnipathy were significantly higher than those in control group (p<0.01). Increased transcription levels of DLG4, Arc and Erg1, and decreased transcription levels of Rab11 and Creb3 were found in patients with somnipathy than in patients without somnipathy. Transcription level of DLG4 was positively, and Rab11 was negatively correlated with their methylation levels. Transcription levels of Arc and Erg1 were positively, and transcription level of Creb3 was negatively correlated with hydroxymethylation levels.</p>
<p>Conclusion: Increased methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 were related to the development of type 2 diabetes in patients with somnipathy. Methylation and hydroxymethylation can significantly affect gene expression at transcription level.</p>',
'date' => '2017-09-03',
'pmid' => 'http://webcache.googleusercontent.com/search?q=cache:t-cxqi84UCcJ:www.alliedacademies.org/articles/transcription-of-synaptic-plasticityrelated-genes-in-patients-with-somnipathy-combined-with-type-2-diabetes.pdf+&cd=1&hl=en&ct=clnk&gl=us',
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'id' => '3265',
'name' => 'Emerging Role of One-Carbon Metabolism and DNA Methylation Enrichment on δ-Containing GABAA Receptor Expression in the Cerebellum of Subjects with Alcohol Use Disorders (AUD',
'authors' => 'Gatta E. et al.',
'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
'date' => '2017-08-19',
'pmid' => 'https://academic.oup.com/ijnp/article/doi/10.1093/ijnp/pyx075/4085582/Emerging-role-of-one-carbon-metabolism-and-DNA',
'doi' => '',
'modified' => '2017-10-09 16:11:05',
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'name' => 'Coordinate Regulation of TET2 and EBNA2 Control DNA Methylation State of Latent Epstein-Barr Virus',
'authors' => 'Lu F. et al.',
'description' => '<p>Epstein-Barr Virus (EBV) latency and its associated carcinogenesis are regulated by dynamic changes in DNA methylation of both virus and host genomes. We show here that the Ten-Eleven Translocation 2 (TET2) gene, implicated in hydroxymethylation and active DNA demethylation, is a key regulator of EBV latency type DNA methylation patterning. EBV latency types are defined by DNA methylation patterns that restrict expression of viral latency genes. We show that TET2 mRNA and protein expression correlate with the highly demethylated EBV type III latency program permissive for expression of EBNA2, EBNA3s, and LMP transcripts. We show that shRNA depletion of TET2 results in a decrease in latency gene expression, but can also trigger a switch to lytic gene expression. TET2 depletion results in the loss of hydroxymethylated cytosine, and corresponding increase in cytosine methylation at key regulatory regions on the viral and host genomes. This also corresponded to a loss of RBP-jκ binding, and decreased histone H3K4 trimethylation at these sites. Furthermore, we show that the TET2 gene, itself, is regulated similar to the EBV genome. ChIP-Seq revealed that TET2 gene contains EBNA2-dependent RBP-jκ and EBF1 binding sites, and is subject to DNA methylation associated transcriptional silencing similar to EBV latency type III genomes. Finally, we provide evidence that TET2 colocalizes with EBNA2-EBF1-RBP-jκ binding sites, and can interact with EBNA2 by co-immunoprecipitation. Taken together, these findings indicate that TET2 gene transcripts are regulated similarly to EBV type III latency genes, and that TET2 protein is a cofactor of EBNA2 and co-regulator of the EBV type III latency program and DNA methylation state..<b>IMPORTANCE</b> Epstein-Barr Virus (EBV) latency and carcinogenesis involves the selective epigenetic modification of viral and cellular genes. Here, we show that TET2, a cellular tumor suppressor involved in active DNA demethylation, plays a central role in regulating DNA methylation state during EBV latency. TET2 is coordinately regulated and functionally interacts with the viral oncogene EBNA2. TET2 and EBNA2 function cooperatively to demethylate genes important for EBV-driven B cells growth transformation.</p>',
'date' => '2017-08-07',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28794029',
'doi' => '',
'modified' => '2017-09-26 09:54:39',
'created' => '2017-09-26 09:54:39',
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'name' => 'Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer',
'authors' => 'Vafadar-Isfahani N. et al.',
'description' => '<p>Hypomethylation of LINE-1 repeats in cancer has been proposed as the main mechanism behind their activation; this assumption, however, was based on findings from early studies that were biased toward young and transpositionally active elements. Here, we investigate the relationship between methylation of 2 intergenic, transpositionally inactive LINE-1 elements and expression of the LINE-1 chimeric transcript (LCT) 13 and LCT14 driven by their antisense promoters (L1-ASP). Our data from DNA modification, expression, and 5'RACE analyses suggest that colorectal cancer methylation in the regions analyzed is not always associated with LCT repression. Consistent with this, in HCT116 colorectal cancer cells lacking DNA methyltransferases DNMT1 or DNMT3B, LCT13 expression decreases, while cells lacking both DNMTs or treated with the DNMT inhibitor 5-azacytidine (5-aza) show no change in LCT13 expression. Interestingly, levels of the H4K20me3 histone modification are inversely associated with LCT13 and LCT14 expression. Moreover, at these LINE-1s, H4K20me3 levels rather than DNA methylation seem to be good predictor of their sensitivity to 5-aza treatment. Therefore, by studying individual LINE-1 promoters we have shown that in some cases these promoters can be active without losing methylation; in addition, we provide evidence that other factors (e.g., H4K20me3 levels) play prominent roles in their regulation.</p>',
'date' => '2017-03-16',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28300471',
'doi' => '',
'modified' => '2017-05-10 16:26:24',
'created' => '2017-05-10 16:26:24',
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(int) 7 => array(
'id' => '2951',
'name' => 'Maternal immune activation induces GAD1 and GAD2 promoter remodeling in the offspring prefrontal cortex',
'authors' => 'Labouesse MA et al.',
'description' => '<p>Maternal infection during pregnancy increases the risk of neurodevelopmental disorders in the offspring. In addition to its influence on other neuronal systems, this early-life environmental adversity has been shown to negatively affect cortical γ-aminobutyric acid (GABA) functions in adult life, including impaired prefrontal expression of enzymes required for GABA synthesis. The underlying molecular processes, however, remain largely unknown. In the present study, we explored whether epigenetic modifications represent a mechanism whereby maternal infection during pregnancy can induce such GABAergic impairments in the offspring. We used an established mouse model of prenatal immune challenge that is based on maternal treatment with the viral mimetic poly(I:C). We found that prenatal immune activation increased prefrontal levels of 5-methylated cytosines (5mC) and 5-hydroxymethylated cytosines (5hmC) in the promoter region of GAD1, which encodes the 67-kDa isoform of the GABA-synthesising enzyme glutamic acid decarboxylase (GAD67). The early-life challenge also increased 5mC levels at the promoter region of GAD2, which encodes the 65-kDa GAD isoform (GAD65). These effects were accompanied by elevated GAD1 and GAD2 promoter binding of methyl CpG-binding protein 2 (MeCP2) and by reduced GAD67 and GAD65 mRNA expression. Moreover, the epigenetic modifications at the GAD1 promoter correlated with prenatal infection-induced impairments in working memory and social interaction. Our study thus highlights that hypermethylation of GAD1 and GAD2 promoters may be an important molecular mechanism linking prenatal infection to presynaptic GABAergic impairments and associated behavioral and cognitive abnormalities in the offspring.</p>',
'date' => '2015-12-02',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26575259',
'doi' => ' 10.1080/15592294.2015.1114202',
'modified' => '2016-06-10 16:32:32',
'created' => '2016-06-10 16:32:32',
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'name' => 'Hepatic DNA hydroxymethylation is site-specifically altered by chronic alcohol consumption and aging',
'authors' => 'Tammen SA et al.',
'description' => '<div class="">
<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26578530',
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'
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'description' => '<p>The DNA hydroxymethylation control package includes one hydroxymethylated spike-in control and its corresponding qPCR primer set that can be added to the DNA sample of interest for any hydroxymethylation profiling experiment (e.g. with Diagenode's <a href="https://www.diagenode.com/en/p/hmedip-kit-x16-monoclonal-mouse-antibody-16-rxns">hMeDIP Kit</a>).</p>
<p><em><strong>CAUTION:</strong> The hydroxymethylated spike-in control is produced from a genomic sequence from Arabidopsis thaliana and may therefore interfere with plant samples. However, it does not show significant homology with other samples species (e.g. human, mouse or rat).</em></p>',
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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<center>
<h4>High DNA recovery after purification of ChIP samples using IPure technology</h4>
<center><img src="https://www.diagenode.com/img/product/kits/ipure-chromatin-function.png" width="500" /></center>
<p></p>
<p><small>ChIP assays were performed using different amounts of U2OS cells and the H3K9me3 antibody (Cat. No.<span> </span><span>C15410056</span>; 2 g/IP). <span>The purified DNA was eluted in 50 µl of water and quantified with a Nanodrop.</span></small></p>
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<li style="text-align: left;">Optimized for DNA purification after ChIP, MeDIP and CUT&Tag</li>
<li style="text-align: left;">Compatible with automation</li>
<li style="text-align: left;">Validated on the IP-Star Compact</li>
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<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-A.png" alt="ChIP-seq figure A" style="display: block; margin-left: auto; margin-right: auto;" /></p>
<p><img src="https://www.diagenode.com/img/product/kits/ideal-TF-chip-seq-B.png" alt="ChIP-seq figure B" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<p><small><strong>Figure 1.</strong> Chromatin Immunoprecipitation has been performed using chromatin from HeLa cells, the iDeal ChIP-seq kit for Transcription Factors (containing the IPure module for DNA purification) and the Diagenode ChIP-seq-grade HDAC1 (A), LSD1 (B) and p53 antibody (C). The IP'd DNA was subsequently analysed on an Illumina® Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. This figure shows the peak distribution in regions of chromosome 3 (A), chromosome 12 (B) and chromosome 6 (C) respectively.</small></p>
<p></p>
<h2>IPure after CUT&Tag</h2>
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<center><img src="https://www.diagenode.com/img/product/kits/ipure-fig2.png" style="display: block; margin-left: auto; margin-right: auto;" width="400" /></center><center>
<p style="text-align: center;"><small><strong>Figure 2.</strong> Heatmap 3kb upstream and downstream of the TSS for H3K4me3</small></p>
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<p></p>
<p><img src="https://www.diagenode.com/img/product/kits/ipure-fig3.png" style="display: block; margin-left: auto; margin-right: auto;" width="600" /></p>
<p></p>
<center><small><strong>Figure 3.</strong> Integrative genomics viewer (IGV) visualization of CUT&Tag experiments using Diagenode’s pA-Tn5 transposase (Cat. No. C01070002), H3K27me3 antibody (Cat. No. C15410069) and IPure kit v2 vs phenol chloroform purification (PC).</small></center>
<p></p>
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<h2>IPure after MeDIP</h2>
<center><img src="https://www.diagenode.com/img/product/kits/magmedip-seq-figure_multi3.jpg" alt="medip sequencing coverage" width="600" /></center><center></center><center>
<p></p>
<small><strong>Figure 4.</strong> Consistent coverage and methylation detection from different starting amounts of DNA with the Diagenode MagMeDIP-seq Package (including the Ipure kit for DNA purification). Samples containing decreasing starting amounts of DNA (from the top down: 1000 ng (red), 250 ng (blue), 100 ng (green)) originating from human blood were prepared, revealing a consistent coverage profile for the three different starting amounts, which enables reproducible methylation detection. The CpG islands (CGIs) (marked by yellow boxes in the bottom track) are predominantly unmethylated in the human genome, and as expected, we see a depletion of reads at and around CGIs.</small></center>
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<h3><strong>Workflow description</strong></h3>
<h5><strong>IPure after ChIP</strong></h5>
<p><strong>Step 1:</strong> Chromatin is decrosslinked and eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added.<br /> <strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet.<br /> <strong>Step 3:</strong> Proteins and remaining buffer are washed away.<br /> <strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after MeDIP</strong></h5>
<p><strong>Step 1:</strong> DNA is eluted from beads (magnetic or agarose) which are discarded. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Remaining buffer are washed away.<br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).<br /><br /><br /></p>
<h5><strong>IPure after CUT&Tag</strong></h5>
<p><strong>Step 1:</strong> pA-Tn5 is inactivated and DNA released from the cells. <strong>Magnetic beads</strong> <strong>for purification</strong> are added. <br /><strong>Step 2:</strong> Magnetic beads acquire positive charge to bind the negatively charged phosphate backbone of DNA. DNA-bead complex is separated using a magnet. <br /><strong>Step 3:</strong> Proteins and remaining buffer are washed away. <br /><strong>Step 4:</strong> DNA is eluted from magnetic beads, which are discarded. Purified DNA is ready for any downstream application (NGS, qPCR, amplification, microarray).</p>
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'slug' => 'ipure-kit-v2-x100',
'meta_title' => 'IPure kit v2 | Diagenode',
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'meta_description' => 'IPure kit v2',
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'name' => '5-hydroxymethylcytosine (5-hmC) Antibody (mouse) ',
'description' => '<p>One of the <strong>only two monoclonal antibodies raised against 5-hydroxymethylcytosine (5-hmC).</strong> 5-hmC is a recently discovered DNA modification which results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. Preliminary results indicate that 5-hmC may have important roles distinct from 5-methylcytosine (5-mC). Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics.</p>
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<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig1.png" alt="ChIP" width="160" caption="false" height="280" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<p><small><strong> Figure 1. An hydroxymethylated DNA IP (hMeDIP) was performed using the Diagenode mouse monoclonal antibody directed against 5-hydroxymethylcytosine (Cat. No. MAb-31HMC-020, MAb-31HMC-050, MAb-31HMC-100).</strong> <br />The IgG isotype antibodies from mouse (Cat. No. kch-819-015) was used as negative control. The DNA was prepared with the GenDNA module of the hMeDIP kit and sonicated with our Bioruptor® (UCD-200/300 series) to have DNA fragments of 300-500 bp. 1 μg of human Hela cells DNA were spiked with non-methylated, methylated, and hydroxymethylated PCR fragments. The IP’d material has been analysed by qPCR using the primer pair specific for the 3 different control sequences. The obtained results show that the mouse monoclonal for 5-hmC is highly specific for this base modification (no IP with non-methylated or methylated C bases containing fragments). </small></p>
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<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig2.png" alt="ELISA" width="190" caption="false" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<p><small><strong> Figure 2. Determination of the 5-hmC mouse monoclonal antibody titer </strong><br />To determine the titer, an ELISA was performed using a serial dilution of the Diagenode mouse monoclonal antibody directed against 5-hmC (Cat No. MAb-31HMC-050, MAb-31HMC-100) in antigen coated wells. The antigen used was KHL coupled to 5-hmC base. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:40,000. </small></p>
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<p><img src="https://www.diagenode.com/img/product/antibodies/C15200200-fig3.png" alt="Dot Blot" width="100" caption="false" height="137" style="display: block; margin-left: auto; margin-right: auto;" /></p>
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<div class="small-8 columns">
<p><small><strong> Figure 3. Dotblot analysis of the Diagenode 5-hmC mouse monoclonal antibody with the C, mC and hmC PCR controls </strong><br />200 to 2 ng (equivalent of 10 to 0.1 pmol of C-bases) of the hmC (1), mC (2) and C (3) PCR controls from the Diagenode “5-hmC, 5-mC & cytosine DNA Standard Pack” (Cat No. AF-101-0020) were spotted on a membrane (Amersham Hybond-N+). The membrane was incubated with 2 μg/ml of the mouse 5-hydroxymethylcytosine monoclonal antibody (dilution 1:500). The membranes were exposed for 30 seconds. </small></p>
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'info2' => '<p>5-hydroxymethylcytosine (5-hmC) has been recently discovered in mammalian DNA. This results from the enzymatic conversion of 5-methylcytosine into 5-hydroxymethylcytosine by the TET family of oxygenases. So far, the 5-hmC bases have been identified in Purkinje neurons, in granule cells and embryonic stem cells where they are present at high levels (up to 0,6% of total nucleotides in Purkinje cells).</p>
<p>Preliminary results indicate that 5-hmC may have important roles distinct from 5-mC. Although its precise role has still to be shown, early evidence suggests a few putative mechanisms that could have big implications in epigenetics : 5-hydroxymethylcytosine may well represent a new pathway to demethylate DNA involving a repair mechanism converting 5-hmC to cytosine and, as such open up entirely new perspectives in epigenetic studies.</p>
<p>Due to the structural similarity between 5-mC and 5-hmC, these bases are experimentally almost indistinguishable. Recent articles demonstrated that the most common approaches (e.g. enzymatic approaches, bisulfite sequencing) do not account for 5-hmC. The development of the affinity-based technologies appears to be the most powerful way to differentially and specifically enrich 5-mC and 5-hmC sequences. The results shown here illustrate the use of this unique monoclonal antibody against 5-hydroxymethylcytosine that has been fully validated in various technologies.</p>',
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'meta_title' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) | Diagenode',
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'meta_description' => '5-hydroxymethylcytosine (5-hmC) Monoclonal Antibody (mouse) validated in hMeDIP, DB and ELISA. Batch-specific data available on the website. Sample size available.',
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'name' => 'DNA hydroxymethylation control package',
'description' => '<p>The DNA hydroxymethylation control package includes one hydroxymethylated spike-in control and its corresponding qPCR primer set that can be added to the DNA sample of interest for any hydroxymethylation profiling experiment (e.g. with Diagenode's <a href="https://www.diagenode.com/en/p/hmedip-kit-x16-monoclonal-mouse-antibody-16-rxns">hMeDIP Kit</a>).</p>
<p><em><strong>CAUTION:</strong> The hydroxymethylated spike-in control is produced from a genomic sequence from Arabidopsis thaliana and may therefore interfere with plant samples. However, it does not show significant homology with other samples species (e.g. human, mouse or rat).</em></p>',
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<div class="small-12 medium-3 large-3 columns"><center><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank"><img src="https://www.diagenode.com/img/banners/banner-nature-publication-580.png" /></a></center></div>
<div class="small-12 medium-9 large-9 columns">
<h3>Sensitive tumour detection and classification using plasma cell-free DNA methylomes<br /><a href="https://www.ncbi.nlm.nih.gov/pubmed/30429608" target="_blank">Read the publication</a></h3>
<h3 class="c-article-title u-h1" data-test="article-title" itemprop="name headline">Preparation of cfMeDIP-seq libraries for methylome profiling of plasma cell-free DNA<br /><a href="https://www.nature.com/articles/s41596-019-0202-2" target="_blank" title="cfMeDIP-seq Nature Method">Read the method</a></h3>
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<div class="row">
<div class="large-12 columns"><span>The Methylated DNA Immunoprecipitation is based on the affinity purification of methylated and hydroxymethylated DNA using, respectively, an antibody directed against 5-methylcytosine (5-mC) in the case of MeDIP or 5-hydroxymethylcytosine (5-hmC) in the case of hMeDIP.</span><br />
<h2></h2>
<h2>How it works</h2>
<p>In brief, Methyl DNA IP is performed as follows: Genomic DNA from cultured cells or tissues is prepared, sheared, and then denatured. Then, immunoselection and immunoprecipitation can take place using the antibody directed against 5 methylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d methylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or next generation sequencing.</p>
<h2>Applications</h2>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-kit-x48-48-rxns" class="center alert radius button"> qPCR analysis</a></div>
<div align="center"><a href="https://www.diagenode.com/en/p/magmedip-seq-package-V2-x10" class="center alert radius button"> NGS analysis </a></div>
<h2>Advantages</h2>
<ul style="font-size: 19px;" class="nobullet">
<li><i class="fa fa-arrow-circle-right"></i> <strong>Unaffected</strong> DNA</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>High enrichment</strong> yield</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>Robust</strong> & <strong>reproducible</strong> techniques</li>
<li><i class="fa fa-arrow-circle-right"></i> <strong>NGS</strong> compatible</li>
</ul>
<h2></h2>
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'meta_description' => 'Methylated DNA immunoprecipitation method is based on the affinity purification of methylated DNA using an antibody directed against 5 methylcytosine (5-mC). ',
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<div class="large-12 columns">
<div style="text-align: justify;" class="small-12 medium-8 large-8 columns">
<h2>Complete solutions for DNA methylation studies</h2>
<p>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements.</p>
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<div class="small-12 medium-4 large-4 columns text-center"><a href="../landing-pages/dna-methylation-grant-applications"><img src="https://www.diagenode.com/img/banners/banner-dna-grant.png" alt="" /></a></div>
<div style="text-align: justify;" class="small-12 medium-12 large-12 columns">
<p>DNA methylation was the first discovered epigenetic mark and is the most widely studied topic in epigenetics. <em>In vivo</em>, DNA is methylated following DNA replication and is involved in a number of biological processes including the regulation of imprinted genes, X chromosome inactivation. and tumor suppressor gene silencing in cancer cells. Methylation often occurs in cytosine-guanine rich regions of DNA (CpG islands), which are commonly upstream of promoter regions.</p>
</div>
<div class="small-12 medium-12 large-12 columns"><br /><br />
<ul class="accordion" data-accordion="">
<li class="accordion-navigation"><a href="#dnamethyl"><i class="fa fa-caret-right"></i> Learn more</a>
<div id="dnamethyl" class="content">5-methylcytosine (5-mC) has been known for a long time as the only modification of DNA for epigenetic regulation. In 2009, however, Kriaucionis discovered a second methylated cytosine, 5-hydroxymethylcytosine (5-hmC). The so-called 6th base, is generated by enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine by the TET family of oxygenases. Early reports suggested that 5-hmC may represent an intermediate of active demethylation in a new pathway which demethylates DNA, converting 5-mC to cytosine. Recent evidence fuel this hypothesis suggesting that further oxidation of the hydroxymethyl group leads to a formyl or carboxyl group followed by either deformylation or decarboxylation. The formyl and carboxyl groups of 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC) could be enzymatically removed without excision of the base.
<p class="text-center"><img src="https://www.diagenode.com/img/categories/kits_dna/dna_methylation_variants.jpg" /></p>
</div>
</li>
</ul>
<br />
<h2>Main DNA methylation technologies</h2>
<p style="text-align: justify;">Overview of the <span style="font-weight: 400;">three main approaches for studying DNA methylation.</span></p>
<div class="row">
<ol>
<li style="font-weight: 400;"><span style="font-weight: 400;">Chemical modification with bisulfite – Bisulfite conversion</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Enrichment of methylated DNA (including MeDIP and MBD)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Treatment with methylation-sensitive or dependent restriction enzymes</span></li>
</ol>
<p><span style="font-weight: 400;"> </span></p>
<div class="row">
<table>
<thead>
<tr>
<th></th>
<th>Description</th>
<th width="350">Features</th>
</tr>
</thead>
<tbody>
<tr>
<td><strong>Bisulfite conversion</strong></td>
<td><span style="font-weight: 400;">Chemical conversion of unmethylated cytosine to uracil. Methylated cytosines are protected from this conversion allowing to determine DNA methylation at single nucleotide resolution.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Single nucleotide resolution</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Quantitative analysis - methylation rate (%)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Gold standard and well studied</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><b>Methylated DNA enrichment</b></td>
<td><span style="font-weight: 400;">(Hydroxy-)Methylated DNA is enriched by using specific antibodies (hMeDIP or MeDIP) or proteins (MBD) that specifically bind methylated CpG sites in fragmented genomic DNA.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Resolution depends on the fragment size of the enriched methylated DNA (300 bp)</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Qualitative analysis</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Compatible with automation</span></li>
</ul>
</td>
</tr>
<tr>
<td><strong>Restriction enzyme-based digestion</strong></td>
<td><span style="font-weight: 400;">Use of (hydroxy)methylation-sensitive or (hydroxy)methylation-dependent restriction enzymes for DNA methylation analysis at specific sites.</span></td>
<td>
<ul style="list-style-type: circle;">
<li style="font-weight: 400;"><span style="font-weight: 400;">Determination of methylation status is limited by the enzyme recognition site</span></li>
<li style="font-weight: 400;"><span style="font-weight: 400;">Easy to use</span></li>
</ul>
</td>
</tr>
</tbody>
</table>
</div>
</div>
<div class="row"></div>
</div>
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<div class="row">
<div class="large-12 columns">The hydroxymethylated DNA IP (hMeDIP) is based on the affinity purification of methylated DNA using an antibody directed against 5-hydroxymethylcytosine (5-hmC).
<h3>How it works</h3>
In brief, hydroxymethyl DNA IP is performed as follows: starting from sheared genomic DNA from cultured cells or tissues, the immunoselection and immunoprecipitation can take place using the antibody directed against 5-hydroxymethylcytosine and antibody binding beads. After isolation and purification is performed, the IP’d hydroxymethylated DNA is ready for any subsequent analysis as qPCR, amplification, hybridization on microarrays or Next Generation Sequencing.
<h3>Overview</h3>
<p class="text-center"><img src="https://www.diagenode.com/img/applications/magnetic_medip_overview.jpg" caption="false" width="726" height="916" /></p>
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<p><span>The Diagenode hMeDIP kit is designed to immunoprecipitate hydroxymethylated DNA (hMethyl DNA IP). This kit is the first and only example of MeDIP kit specifically designed and fully validated for affinity-capture and detection of hydroxymethylated regions using the highly specific rat, mouse or rabbit antibodies against 5-hmC. </span></p>
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'description' => '<p>Cytosine hydroxymethylation was recently discovered as an important epigenetic mechanism. This cytosine base modification results from the enzymatic conversion of 5-methylcytosine (5-mC) into 5-hydroxymethylcytosine (5-hmC) by the TET family of oxygenases. Though the precise role of 5-hmC is the subject of intense research and debate, early studies strongly indicate that it is also involved in gene regulation and in numerous important biological processes including embryonic development, cellular differentiation, stem cell reprogramming and carcinogenesis.</p>
<p>The study of 5-hmC has long been limited due to the lack of high quality, validated tools and technologies that discriminate hydroxymethylation from methylation in regulating gene expression. The use of highly specific antibodies against 5-hmC for the immunoprecipitation of hydroxymethylated DNA offers a reliable solution for hydroxymethylation profiling.</p>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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'id' => '1778',
'name' => 'product/kits/methyl-kit-icon.png',
'alt' => 'Methylation kit icon',
'modified' => '2019-04-23 15:17:01',
'created' => '2018-03-15 15:52:12',
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'id' => '4244',
'name' => 'Developmental and Injury-induced Changes in DNA Methylation inRegenerative versus Non-regenerative Regions of the VertebrateCentral Nervous System',
'authors' => 'Reverdatto S. et al.',
'description' => '<p>Background Because some of its CNS neurons (e.g., retinal ganglion cells after optic nerve crush (ONC)) regenerate axons throughout life, whereas others (e.g., hindbrain neurons after spinal cord injury (SCI)) lose this capacity as tadpoles metamorphose into frogs, the South African claw-toed frog, Xenopus laevis, offers unique opportunities for exploring differences between regenerative and non-regenerative responses to CNS injury within the same organism. An earlier, three-way RNA-seq study (frog ONC eye, tadpole SCI hindbrain, frog SCI hindbrain) identified genes that regulate chromatin accessibility among those that were differentially expressed in regenerative vs non-regenerative CNS [11]. The current study used whole genome bisulfite sequencing (WGBS) of DNA collected from these same animals at the peak period of axon regeneration to study the extent to which DNA methylation could potentially underlie differences in chromatin accessibility between regenerative and non-regenerative CNS. Results Consistent with the hypothesis that DNA of regenerative CNS is more accessible than that of non-regenerative CNS, DNA from both the regenerative tadpole hindbrain and frog eye was less methylated than that of the non-regenerative frog hindbrain. Also, consistent with observations of CNS injury in mammals, DNA methylation in non-regenerative frog hindbrain decreased after SCI. However, contrary to expectations that the level of DNA methylation would decrease even further with axotomy in regenerative CNS, DNA methylation in these regions instead increased with injury. Injury-induced differences in CpG methylation in regenerative CNS became especially enriched in gene promoter regions, whereas non-CpG methylation differences were more evenly distributed across promoter regions, intergenic, and intragenic regions. In non-regenerative CNS, tissue-related (i.e., regenerative vs. non-regenerative CNS) and injury-induced decreases in promoter region CpG methylation were significantly correlated with increased RNA expression, but the injury-induced, increased CpG methylation seen in regenerative CNS across promoter regions was not, suggesting it was associated with increased rather than decreased chromatin accessibility. This hypothesis received support from observations that in regenerative CNS, many genes exhibiting increased, injury-induced, promoter-associated CpG-methylation also exhibited increased RNA expression and association with histone markers for active promoters and enhancers. DNA immunoprecipitation for 5hmC in optic nerve regeneration found that the promoter-associated increases seen in CpG methylation were distinct from those exhibiting changes in 5hmC. Conclusions Although seemingly paradoxical, the increased injury-associated DNA methylation seen in regenerative CNS has many parallels in stem cells and cancer. Thus, these axotomy-induced changes in DNA methylation in regenerative CNS provide evidence for a novel epigenetic state favoring successful over unsuccessful CNS axon regeneration. The datasets described in this study should help lay the foundations for future studies of the molecular and cellular mechanisms involved. The insights gained should, in turn, help point the way to novel therapeutic approaches for treating CNS injury in mammals. Supplementary Information The online version contains supplementary material available at 10.1186/s12864-021-08247-0.</p>',
'date' => '2022-01-01',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/34979916',
'doi' => '10.1186/s12864-021-08247-0',
'modified' => '2022-05-20 09:20:25',
'created' => '2022-05-19 10:41:50',
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'id' => '4271',
'name' => 'Bone marrow age dictates clonality of smooth muscle-derived cells in theatherosclerotic plaque',
'authors' => 'Kabir Inamul et al.',
'description' => '<p>Aging is the predominant risk factor for atherosclerosis, the leading cause of death. Rare smooth muscle cells (SMCs) clonally expand giving rise to up to ∼70\% of atherosclerotic plaque cells; however, the effect of age on SMC clonality is not known. Our results indicate that aging induces SMC polyclonality and worsens atherosclerosis through non-cell autonomous effects of aged bone marrow-derived cells. Indeed, in myeloid cells from aged mice and humans, TET2 levels are reduced which epigenetically silences integrin β3 resulting in increased cytokine (e.g., tumor necrosis factor [TNF]-α) signaling. In turn, TNFα induces recruitment and expansion of multiple SMCs into the atherosclerotic plaque. Recent studies demonstrate that normal aging is characterized by somatic mutations and clonal expansion of epithelial cells of diverse tissues (e.g., esophagus, endometrium, skin); extrapolating beyond atherogenesis, our results call for future studies evaluating the role of aged myeloid cells in regulating this epithelial cell clonal expansion.</p>',
'date' => '2022-01-01',
'pmid' => 'https://doi.org/10.1101%2F2022.01.18.476756',
'doi' => '10.1101/2022.01.18.476756',
'modified' => '2022-05-23 09:45:53',
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'id' => '4329',
'name' => 'Epigenetic remodelling of enhancers in response to estrogen deprivationand re-stimulation.',
'authors' => 'Sklias Athena et al.',
'description' => '<p>Estrogen hormones are implicated in a majority of breast cancers and estrogen receptor alpha (ER), the main nuclear factor mediating estrogen signaling, orchestrates a complex molecular circuitry that is not yet fully elucidated. Here, we investigated genome-wide DNA methylation, histone acetylation and transcription after estradiol (E2) deprivation and re-stimulation to better characterize the ability of ER to coordinate gene regulation. We found that E2 deprivation mostly resulted in DNA hypermethylation and histone deacetylation in enhancers. Transcriptome analysis revealed that E2 deprivation leads to a global down-regulation in gene expression, and more specifically of TET2 demethylase that may be involved in the DNA hypermethylation following short-term E2 deprivation. Further enrichment analysis of transcription factor (TF) binding and motif occurrence highlights the importance of ER connection mainly with two partner TF families, AP-1 and FOX. These interactions take place in the proximity of E2 deprivation-mediated differentially methylated and histone acetylated enhancers. Finally, while most deprivation-dependent epigenetic changes were reversed following E2 re-stimulation, DNA hypermethylation and H3K27 deacetylation at certain enhancers were partially retained. Overall, these results show that inactivation of ER mediates rapid and mostly reversible epigenetic changes at enhancers, and bring new insight into early events, which may ultimately lead to endocrine resistance.</p>',
'date' => '2021-09-01',
'pmid' => 'https://doi.org/10.1093%2Fnar%2Fgkab697',
'doi' => '10.1093/nar/gkab697',
'modified' => '2022-06-22 09:25:09',
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'id' => '3245',
'name' => 'Transcription of synaptic plasticity-related genes in patients with somnipathy combined with type 2 diabetes',
'authors' => 'Yi Zhang, Rui Ma, Shaohong Zou, Gaiyu Tong, Gulibakeranmu Abula, Manna Hu, Qing Dai',
'description' => '<p>Objective: To investigate DNA methylation and hydroxymethylation in patients with somnipathy combined with type 2 diabetes, and to explore the effects of DNA methylation and hydroxymethylation on gene expression.</p>
<p>Methods: Thirty patients with somnipathy combined with type 2 diabetes and 20 patients with type-2 diabetes but without somnipathy were considered. DNA methylation of Disks Large Homolog 4 (DLG4) and Ras-related protein Rab-11 (Rab11) was detected by bisulfite sequencing and DNA hydroxymethylation of activity-regulated cytoskeleton-associated protein (Arc), Cyclic AMP-Responsive Element-Binding protein 3 (CREB3) and Early Growth Response protein 1 (EGR1) was analyzed by CHIP analysis. Transcription levels of DLG4, Rab11, Arc, CREB3 and EGR1 were detected by quantitative real-time RT-PCR (qRT-PCR).</p>
<p>Results: Methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 in patients with somnipathy were significantly higher than those in control group (p<0.01). Increased transcription levels of DLG4, Arc and Erg1, and decreased transcription levels of Rab11 and Creb3 were found in patients with somnipathy than in patients without somnipathy. Transcription level of DLG4 was positively, and Rab11 was negatively correlated with their methylation levels. Transcription levels of Arc and Erg1 were positively, and transcription level of Creb3 was negatively correlated with hydroxymethylation levels.</p>
<p>Conclusion: Increased methylation levels of DLG4 and Rab11 and hydroxymethylation levels of Arc, Creb3 and Erg1 were related to the development of type 2 diabetes in patients with somnipathy. Methylation and hydroxymethylation can significantly affect gene expression at transcription level.</p>',
'date' => '2017-09-03',
'pmid' => 'http://webcache.googleusercontent.com/search?q=cache:t-cxqi84UCcJ:www.alliedacademies.org/articles/transcription-of-synaptic-plasticityrelated-genes-in-patients-with-somnipathy-combined-with-type-2-diabetes.pdf+&cd=1&hl=en&ct=clnk&gl=us',
'doi' => '',
'modified' => '2017-09-25 08:51:27',
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'id' => '3265',
'name' => 'Emerging Role of One-Carbon Metabolism and DNA Methylation Enrichment on δ-Containing GABAA Receptor Expression in the Cerebellum of Subjects with Alcohol Use Disorders (AUD',
'authors' => 'Gatta E. et al.',
'description' => '<section class="abstract">
<section class="sec">
<div class="title -title">Background</div>
<p>Cerebellum is an area of the brain particularly sensitive to the effects of acute and chronic alcohol consumption. Alcohol exposure decreases cerebellar Purkinje cell output by increasing GABA release from Golgi cells onto extrasynaptic α<sub>6</sub>/δ-containing GABA<sub>A</sub> receptors located on glutamatergic granule cells. Here, we studied whether chronic alcohol consumption induces changes in GABA<sub>A</sub> receptor subunit expression and whether these changes are associated with alterations in epigenetic mechanisms via DNA methylation.</p>
</section>
<section class="sec">
<div class="title -title">Methods</div>
<p>We used a cohort of postmortem cerebellum from control and chronic alcoholics, here defined as alcohol use disorders subjects (n=25/group). <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine were measured by high-performance liquid chromatography. mRNA levels of various genes were assessed by reverse transcriptase-quantitative polymerase chain reaction. Promoter methylation enrichment was assessed using methylated DNA immunoprecipitation and hydroxy-methylated DNA immunoprecipitation assays.</p>
</section>
<section class="sec">
<div class="title -title">Results</div>
<p>mRNAs encoding key enzymes of 1-carbon metabolism that determine the <em>S</em>-adenosyl-methionine/<em>S</em>-adenosyl-homocysteine ratio were increased, indicating higher “methylation index” in alcohol use disorder subjects. We found that increased methylation of the promoter of the δ subunit GABA<sub>A</sub> receptor was associated with reduced mRNA and protein levels in the cerebellum of alcohol use disorder subjects. No changes were observed in α<sub>1</sub>- or α<sub>6</sub>-containing GABA<sub>A</sub> receptor subunits. The expression of DNA-methyltransferases (1, 3A, and 3B) was unaltered, whereas the mRNA level of TET1, which participates in the DNA demethylation pathway, was decreased. Hence, increased methylation of the δ subunit GABA<sub>A</sub> receptor promoter may result from alcohol-induced reduction of DNA demethylation.</p>
</section>
<section class="sec">
<div class="title -title">Conclusion</div>
<p>Together, these results support the hypothesis that aberrant DNA methylation pathways may be involved in cerebellar pathophysiology of alcoholism. Furthermore, this work provides novel evidence for a central role of DNA methylation mechanisms in the alcohol-induced neuroadaptive changes of human cerebellar GABA<sub>A</sub> receptor function.</p>
</section>
</section>',
'date' => '2017-08-19',
'pmid' => 'https://academic.oup.com/ijnp/article/doi/10.1093/ijnp/pyx075/4085582/Emerging-role-of-one-carbon-metabolism-and-DNA',
'doi' => '',
'modified' => '2017-10-09 16:11:05',
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'id' => '3251',
'name' => 'Coordinate Regulation of TET2 and EBNA2 Control DNA Methylation State of Latent Epstein-Barr Virus',
'authors' => 'Lu F. et al.',
'description' => '<p>Epstein-Barr Virus (EBV) latency and its associated carcinogenesis are regulated by dynamic changes in DNA methylation of both virus and host genomes. We show here that the Ten-Eleven Translocation 2 (TET2) gene, implicated in hydroxymethylation and active DNA demethylation, is a key regulator of EBV latency type DNA methylation patterning. EBV latency types are defined by DNA methylation patterns that restrict expression of viral latency genes. We show that TET2 mRNA and protein expression correlate with the highly demethylated EBV type III latency program permissive for expression of EBNA2, EBNA3s, and LMP transcripts. We show that shRNA depletion of TET2 results in a decrease in latency gene expression, but can also trigger a switch to lytic gene expression. TET2 depletion results in the loss of hydroxymethylated cytosine, and corresponding increase in cytosine methylation at key regulatory regions on the viral and host genomes. This also corresponded to a loss of RBP-jκ binding, and decreased histone H3K4 trimethylation at these sites. Furthermore, we show that the TET2 gene, itself, is regulated similar to the EBV genome. ChIP-Seq revealed that TET2 gene contains EBNA2-dependent RBP-jκ and EBF1 binding sites, and is subject to DNA methylation associated transcriptional silencing similar to EBV latency type III genomes. Finally, we provide evidence that TET2 colocalizes with EBNA2-EBF1-RBP-jκ binding sites, and can interact with EBNA2 by co-immunoprecipitation. Taken together, these findings indicate that TET2 gene transcripts are regulated similarly to EBV type III latency genes, and that TET2 protein is a cofactor of EBNA2 and co-regulator of the EBV type III latency program and DNA methylation state..<b>IMPORTANCE</b> Epstein-Barr Virus (EBV) latency and carcinogenesis involves the selective epigenetic modification of viral and cellular genes. Here, we show that TET2, a cellular tumor suppressor involved in active DNA demethylation, plays a central role in regulating DNA methylation state during EBV latency. TET2 is coordinately regulated and functionally interacts with the viral oncogene EBNA2. TET2 and EBNA2 function cooperatively to demethylate genes important for EBV-driven B cells growth transformation.</p>',
'date' => '2017-08-07',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28794029',
'doi' => '',
'modified' => '2017-09-26 09:54:39',
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(int) 6 => array(
'id' => '3172',
'name' => 'Decoupling of DNA methylation and activity of intergenic LINE-1 promoters in colorectal cancer',
'authors' => 'Vafadar-Isfahani N. et al.',
'description' => '<p>Hypomethylation of LINE-1 repeats in cancer has been proposed as the main mechanism behind their activation; this assumption, however, was based on findings from early studies that were biased toward young and transpositionally active elements. Here, we investigate the relationship between methylation of 2 intergenic, transpositionally inactive LINE-1 elements and expression of the LINE-1 chimeric transcript (LCT) 13 and LCT14 driven by their antisense promoters (L1-ASP). Our data from DNA modification, expression, and 5'RACE analyses suggest that colorectal cancer methylation in the regions analyzed is not always associated with LCT repression. Consistent with this, in HCT116 colorectal cancer cells lacking DNA methyltransferases DNMT1 or DNMT3B, LCT13 expression decreases, while cells lacking both DNMTs or treated with the DNMT inhibitor 5-azacytidine (5-aza) show no change in LCT13 expression. Interestingly, levels of the H4K20me3 histone modification are inversely associated with LCT13 and LCT14 expression. Moreover, at these LINE-1s, H4K20me3 levels rather than DNA methylation seem to be good predictor of their sensitivity to 5-aza treatment. Therefore, by studying individual LINE-1 promoters we have shown that in some cases these promoters can be active without losing methylation; in addition, we provide evidence that other factors (e.g., H4K20me3 levels) play prominent roles in their regulation.</p>',
'date' => '2017-03-16',
'pmid' => 'https://www.ncbi.nlm.nih.gov/pubmed/28300471',
'doi' => '',
'modified' => '2017-05-10 16:26:24',
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(int) 7 => array(
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'name' => 'Maternal immune activation induces GAD1 and GAD2 promoter remodeling in the offspring prefrontal cortex',
'authors' => 'Labouesse MA et al.',
'description' => '<p>Maternal infection during pregnancy increases the risk of neurodevelopmental disorders in the offspring. In addition to its influence on other neuronal systems, this early-life environmental adversity has been shown to negatively affect cortical γ-aminobutyric acid (GABA) functions in adult life, including impaired prefrontal expression of enzymes required for GABA synthesis. The underlying molecular processes, however, remain largely unknown. In the present study, we explored whether epigenetic modifications represent a mechanism whereby maternal infection during pregnancy can induce such GABAergic impairments in the offspring. We used an established mouse model of prenatal immune challenge that is based on maternal treatment with the viral mimetic poly(I:C). We found that prenatal immune activation increased prefrontal levels of 5-methylated cytosines (5mC) and 5-hydroxymethylated cytosines (5hmC) in the promoter region of GAD1, which encodes the 67-kDa isoform of the GABA-synthesising enzyme glutamic acid decarboxylase (GAD67). The early-life challenge also increased 5mC levels at the promoter region of GAD2, which encodes the 65-kDa GAD isoform (GAD65). These effects were accompanied by elevated GAD1 and GAD2 promoter binding of methyl CpG-binding protein 2 (MeCP2) and by reduced GAD67 and GAD65 mRNA expression. Moreover, the epigenetic modifications at the GAD1 promoter correlated with prenatal infection-induced impairments in working memory and social interaction. Our study thus highlights that hypermethylation of GAD1 and GAD2 promoters may be an important molecular mechanism linking prenatal infection to presynaptic GABAergic impairments and associated behavioral and cognitive abnormalities in the offspring.</p>',
'date' => '2015-12-02',
'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26575259',
'doi' => ' 10.1080/15592294.2015.1114202',
'modified' => '2016-06-10 16:32:32',
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(int) 8 => array(
'id' => '2950',
'name' => 'Hepatic DNA hydroxymethylation is site-specifically altered by chronic alcohol consumption and aging',
'authors' => 'Tammen SA et al.',
'description' => '<div class="">
<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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<button class="alert small button expand" onclick="$(this).addToCart('IPure kit v2',
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</form><a class="close-reveal-modal" aria-label="Close">×</a></div><!-- END: ADD TO CART MODAL --><a href="#" id="5-hmc-monoclonal-antibody-mouse-classic-50-ug-50-ul" data-reveal-id="cartModal-2009" class="" style="color:#B21329"><i class="fa fa-cart-plus"></i></a>
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'C02040018',
'145',
$('#CartQuantity').val());" name="keepshop" id="keepshop" type="submit">继续购物</button> </div>
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</div>
</div>
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<p><span>Complete solution for DNA methylation studies! </span></p>
<p><span>Whether you are experienced or new to the field of DNA methylation, Diagenode has everything you need to make your assay as easy and convenient as possible while ensuring consistent data between samples and experiments. Diagenode offers sonication instruments, reagent kits, high quality antibodies, and high-throughput automation capability to address all of your specific DNA methylation analysis requirements. </span></p>
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<h4>PURPOSE:</h4>
<p><abstracttext label="PURPOSE" nlmcategory="OBJECTIVE">Global DNA hydroxymethylation is markedly decreased in human cancers, including hepatocellular carcinoma, which is associated with chronic alcohol consumption and aging. Because gene-specific changes in hydroxymethylcytosine may affect gene transcription, giving rise to a carcinogenic environment, we determined genome-wide site-specific changes in hepatic hydroxymethylcytosine that are associated with chronic alcohol consumption and aging.</abstracttext></p>
<h4>METHODS:</h4>
<p><abstracttext label="METHODS" nlmcategory="METHODS">Young (4 months) and old (18 months) male C57Bl/6 mice were fed either an ethanol-containing Lieber-DeCarli liquid diet or an isocaloric control diet for 5 weeks. Genomic and gene-specific hydroxymethylcytosine patterns were determined through hydroxymethyl DNA immunoprecipitation array in hepatic DNA.</abstracttext></p>
<h4>RESULTS:</h4>
<p><abstracttext label="RESULTS" nlmcategory="RESULTS">Hydroxymethylcytosine patterns were more perturbed by alcohol consumption in young mice than in old mice (431 differentially hydroxymethylated regions, DhMRs, in young vs 189 DhMRs in old). A CpG island ~2.5 kb upstream of the glucocorticoid receptor gene, Nr3c1, had increased hydroxymethylation as well as increased mRNA expression (p = 0.015) in young mice fed alcohol relative to the control group. Aging alone also altered hydroxymethylcytosine patterns, with 331 DhMRs, but alcohol attenuated this effect. Aging was associated with a decrease in hydroxymethylcytosine ~1 kb upstream of the leptin receptor gene, Lepr, and decreased transcription of this gene (p = 0.029). Nr3c1 and Lepr are both involved in hepatic lipid homeostasis and hepatosteatosis, which may create a carcinogenic environment.</abstracttext></p>
<h4>CONCLUSIONS:</h4>
<p><abstracttext label="CONCLUSIONS" nlmcategory="CONCLUSIONS">These results suggest that the location of hydroxymethylcytosine in the genome is site specific and not random, and that changes in hydroxymethylation may play a role in the liver's response to aging and alcohol.</abstracttext></p>
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'pmid' => 'http://www.ncbi.nlm.nih.gov/pubmed/26578530',
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[main] - APP/webroot/index.php, line 118
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