Diagenode

CATS Total RNA-seq Kit (with rRNA depletion) v2 x24

カタログ番号
フォーマット
価格
C05010042
24 rxns
$2,545.00
ブローカー情報

Diagenodeの「CATS small RNA-seq Kit」は、革新的なCATS法(Capture and Amplification by Tailing and Switching)を採用することで、ライゲーションが不要でかつ低インプットRNA量から次世代シークエンシングのDNAライブラリーを作製することができるキットです。

  • 多様な転写産物の検出
  • 100pgの超少量インプットRNAで優れた再現性
  • 難しいサンプルでの優れたパフォーマンス
  • 続きを読む

    CATSは、RNAの3’末端をキャプチャーするためのポリヌクレオチドテーリングおよび、cDNA合成時に5’末端をキャプチャーするためのMMLV-RTによるテンプレートスイッチ 能力を利用した技術です。この方法なら、rRNAが 除去済のサンプルを使用の場合、100ピコグラムの低インプットRNA量からわずか数時間でIllumina用シークエンシングライブラリーを作製することができます。

    ライブラリーは、ストランド特異的な情報を保持しています。「CATS RNA-seq Kit」は、増幅を最小限に抑えることでより多くの有効な鋳型をキャプチャーし、また、バイアスを最小限に抑えることで高い複雑性を得ることができるので、他のライゲーションベースの手法と比べより高いライブラリー作製効率を示します。

More transcripts detected

Biotypes* represented in the libraries CATS v2 total RNA-seq Kit vs NEBNext Ultra I directional RNA-seq Kit. TPM ≥2.
Input: 1 ng, 10 ng rRNA(-) RNA from human universal total RNA (Agilent, 740000). * < 2% of total detected transcripts at this threshold is rRNA

Very reproducible library preparation method

Transcripts detected in two technical replicates of CATS v2 total RNA-seq libraries. TPM≥2
Input: 1 ng rRNA(-) RNA from human universal total RNA (Agilent, 740000).

High correlation in coding transcripts detection across different inputs

Transcripts detected in 10 ng vs 1 ng samples of CATS v2 total RNA-seq libraries. TPM≥2
Input: 1 ng, 10 ng rRNA(-) RNA from human universal total RNA (Agilent, 740000).

Few PCR cycles, less amplification bias

BioAnalyzer® electropherogram of two technical replicates of CATS v2 total RNA-seq libraries.
Input: 1 ng rRNA(-) RNA from human universal total RNA (Agilent, 740000).

CATS saves time



  • 特徴

    This kit contains and rRNA depletion module that depletes both cytoplasmic (5S rRNA, 5.8S rRNA, 18S rRNA, 28S rRNA) and mitochondrial ribosomal RNA (12S rRNA and 16S rRNA) from human, mouse and rat total RNA samples. This product is suitable for both intact and degraded RNA preparations (e.g. FFPE tissue). 

    • Ligation-free  assay - increased efficiency and minimum bias
    • Low inputs down to 100 picograms
    • User-friendly: reduced hands-on  time with fewer steps than competing methods
    • Libraries ready to sequence in only 5 hours
    • Retaining strand specificity of origin

    CATS total RNA-seq rRNA (-) library preparation workflow

    Total time Hands-on-time

    CATS total RNA-seq
    rRNA depletion (module included)

    60-120 min
    (1)

    SAFE STORAGE

    STEP 1
    ssRNA end-repair
    and polyA tailing

    75 min 10 min
    (2)

    STEP 2
    RT and template switch

    150 min 10 min
    (3)

    SAFE STORAGE

    STEP 3
    PCR amplification
    of DNA library

    30 min 5 min
    (4)

    SAFE STORAGE

    (1) RNA: stored at -80°C after purification (safe stop & storage) ; (2) cDNA: stable at 4°C overnight, store at -20°C ; (3) Amplified library: prior purification and QC store at -20°C (safe stop & storage) ; (4) DNA libraries: store at -20°C or -80°C (safe stop & storage)

    Schematic representation of the workflow used by the CATS Small RNA-seq Kit. Single stranded RNAs are first dephosphorylated (end-repaired) and polyadenylated at the 3’-end. Subsequently, a cDNA strand synthesis is performed in the presence of the anchored poly(dT) oligonucleotide containing terminal P7 Illumina® adaptor sequence. When the reverse transcriptase reaches the 5’-end of the RNA it switches the template and continue DNA synthesis over the template-switching oligonucleotide (TSO). The TSO contains three 3’-terminal ribonucleotides X (rX) which facilitate the template switching and carry the terminal P5 Illumina® adaptor sequence. During PCR pre-amplification of the first cDNA strand, Illumina® adapters carrying P5 and P7 terminal sequences (required for clustering on an Illumina® flow cell) as well as index sequences are incorporated into the library. The sum size of the adapters (the size of “empty” library) is 143 bp.

  •  プロトコル集
    CATS RNA-seq v2 library preparation protocol on RNA from FFPE-samples
    The following protocol offers a streamlined solution for whole transcriptome sequencing studies f...
    Download
  • FAQs
    • In the CATS mRNA-seq kit, will the natural polyA tails undergo the polyadenylation step?
      Yes. The RNA fragments containing the natural polyA tail will be tailed as well during the polynucleotide tailing step.
    • What fragmentation should I carry if my sample has a RIN of 2-3?
      For heavily degraded RNA samples with a RIN of < 3, no fragmentation step is recommended to start the library preparation. However, the fragmentation buffer still needs to be added in the sample but no heat should be applied (please skip high temperatures in the “fragmentation step” in manuals).
    • How long is the CATS RNA-seq library preparation protocol?
      For the RNA-seq library preparation itself, takes 4h30-5h to complete for a trained user. This does not include the sample preparation (e.g. polyA selection, rRNA depletion or small RNA enrichment) and the QC of the library. Each of those two steps take additional approximate 1h.
    • I am working with mid- to low-quality RNA. Are there any recommendations?
      We recommend to run a BioAnalyzer (or equivalent) electrophoresis to get the fragmentation profile of the RNA. Then you should decide in collaboration with the Diagenode technical support customer.support@diagenode.com about the best fragmentation time to use for your RNA samples.
    • What would be the lowest possible limit for the input for rRNA depletion and polyA selection modules in CATS kits?
      It is possible to use lower amount of starting material than mentioned in manuals. To get more information about this, contact Diagenode technical support.

    • Before mapping, CATS RNA-seq data requires a specific trimming procedure described in all manuals. Please consult also a user guide “Diagenode trimming tools for CATS RNA-seq”.
    • 7) Is CATS compatible with directional mRNA-seq?
      Yes. CATS RNA-seq library preparation retains strand specificity which is also known as directional library preparation.
    • 8) Which RIN do my RNA samples need to have to work with CATS RNA-seq kits?
      We suggest using high quality RNA (RIN>8) but degraded RNA can be also used. In case of degraded RNA, the fragmentation time must be adjusted. In the protocol we advise 7 min at 94°C for fragmenting high quality RNA (RIN>8). If the RNA is severely degraded like RNA derived from FFPE tissue (RIN 2-3), the fragmentation may be skipped.
  •  資料
    CATS Total RNA-seq Kit MANUAL
    CATS Total RNA-seq Kit (with rRNA depletion): CATS Total RNA sequencing kit for Illumin...
    Download
    CATS paired-end sequencing primer (for read2) DATASHEET
    This primer has been specifically designed to be used in paired-end sequencing with CATS librarie...
    Download
    Diagenode trimming tools for CATS library preparation [read1]
    Due to special mechanisms in creating CATS libraries such as template switching and artificial po...
    Download
    Diagenode trimming tools for CATS library preparation [read2]
    Due to special mechanisms in creating CATS libraries such as template switching and artificial po...
    Download
    Diagenode trimming tools for CATS library preparation MANUAL
    This document describes the usage of the Diagenode trimming tools for CATS library preparation, w...
    Download
    Material Safety Data Sheet CATS total RNA-seq (with rRNA depletion) Kit DATASHEET
    Download
  •  出版物

    How to properly cite this product in your work

    Diagenode strongly recommends using this: CATS Total RNA-seq Kit (with rRNA depletion) v2 x24 (Diagenode Cat# C05010042). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Characterization of the salmon louse Lepeophtheirus salmonis miRNome: Sex-biased differences related to the coding and non-coding RNA interplay.
    Núñez-Acuña G, Gallardo-Escárate C
    The salmon louse Lepeophtheirus salmonis is a marine ectoparasite that has a detrimental impact on salmon farms. Genomic knowledge of adult stages is critical to understand the reproductive success and lifecycle completion of this species. Here, we report a comprehensive characterization of the L. salmonis miRNome w...

    The Atlantic salmon (Salmo salar) antimicrobial peptide cathelicidin-2 is a molecular host-associated cue for the salmon louse (Lepeophtheirus salmonis)
    Gustavo Núñez-Acuña, Cristian Gallardo-Escárate, David M. Fields, Steven Shema, Anne Berit Skiftesvik, Ignacio Ormazábal & Howard I. Browman
    Chemical signals are a key element of host-parasite interactions. In marine ecosystems, obligate ectoparasites, such as sea lice, use chemical cues and other sensory signals to increase the probability of encountering a host and to identify appropriate hosts on which they depend to complete their life cycle. The che...

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