Diagenode

NGSライブラリー調製

RNA-seqのライブラリー作製

Diagenode’s new RNA-sequencing solutions utilize the innovative “Capture and Amplification by Tailing and Switching” (CATS), a ligation-free method to produce DNA libraries for next generation sequencing from low input amounts of RNA.

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    CATS relies on polynucleotide tailing for capturing the 3’-ends of nucleic acids and the template switching ability of MMLV-RT to capture the 5’-ends for cDNA synthesis.

    The Diagenode kits generate Illumina compatible ready-to-sequence libraries from RNA inputs as low as 10 pg in just few hours. Libraries retain strand specificity of origin. Unlike competing solutions, our CATS RNA-seq Kits exhibit higher library efficiency versus ligation-based methods. The CATS method allows for accurate identification of a greater number of non-coding RNAs, providing higher complexity from better template capture and minimal bias due to lower amplification requirements.

  • Diverse transcripts detection
  • Excellent reproducibility with ultra low inputs down to 10 pg
  • Great performance on challenging samples

Do you care about time, reproducibility and ease? Read more about CATS mRNA-seq

CATS small RNA-seq on plasma-derived small RNA samples – Read our customer’s testimonial

Broad overview of the transcriptome

Biotypes* represented in the libraries CATS v2 mRNA-seq Kit vs NEBNext Ultra I directional RNA-seq Kit. TPM ≥2.
Input: 1 ng, 10 ng poly(A) RNA from human universal total RNA (Agilent, 740000). * < 0.04% of total detected transcripts at this threshold is rRNA

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

Uniquely detected transcripts

All detected transcripts in CATS v2 mRNA-seq Kit vs NEBNext Ultra I directional library preparation Kit. No threshold has been set for the analysis.
Input: 1 ng poly(A) RNA from human universal total RNA (Agilent, 740000). 36,245 transcripts found in both libraries.

Consistency and wide representation of the transcriptome

Comparison on the number of coding transcripts detected with CATS v2 mRNA-seq Kit vs SMARTer stranded RNA-seq library preparation kit. TPM ≥2.
Input: 1 ng poly(A) RNA from human universal total RNA (Agilent, 740000). 21,333 transcripts found in both libraries.

High-complexity at low inputs

Mapped reads distribution for CATS v2 mRNA-seq Kit vs NEBNext Ultra I directional RNA-seq Kit.
Input: 1 ng, 10 ng poly(A) RNA from human universal total RNA (Agilent, 740000).

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 transcripts detection across ultra low inputs

Transcripts detected in 1 ng vs 0.1 ng samples of CATS v2 mRNA-seq libraries. TPM≥2.
Input: 1 ng, 0.1 ng poly(A) RNA from human universal total RNA (Agilent, 740000).

Excellent performance on challenging samples

Heatmap clustering of the top 50 differentially expressed microRNAs(miRNAs) after library preparation:

  • CATS Small RNA-seq Kit on 1 ng exosomal RNA (Exosome sample 1, 2)
  • CATS RNA-seq Kit on 1 ng total RNA from parent (HEK293T) cells (Cell sample 1, 2).

Prior library preparation, exosomes were isolated with Diagenode exosome capture solutions Exosomes

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).

Ultra low inputs, excellent reproducibility

Correlation between 2 different operators using CATS small RNA-seq Kit on two different inputs (100 pg vs 1 ng) for ncRNA detected at TPM ≥2.
R2 = 0.86

Ultra low inputs, few PCR cycles

Representation of the library preparation yield according to different methods (CATS small RNA-seq Kit vs NEBNext UltraI kit) for 100 ng total RNA. Number of PCR cycles applied following manufacturer’s instructions.

Consistency across template switching techniques

Comparison of the number of detected small non-coding RNAs CATS small RNA-seq Kit vs SMARTer
Input: 100 pg isolated small RNA. TPM ≥2. Transcripts taken into consideration: miRNA, piRNA, scRNA, scaRNA, snoRNA, snRNA and vaultRNA. 1,217 transcripts found in both libraries at this expression level.

CATS saves time



CATS for Small RNA White Paper:

Low input RNA-seq library preparation provides higher small non-coding RNA diversity and greatly reduced hands-on time

Prior to RNA sequencing, the preparation of suitable libraries for the NGS platform is required . The library preparation typically requires the conversion of the template RNA into cDNA and the fusion of adapters to both ends before PCR amplification.For ligation-based techniques, the addition of the adapters by RNA ligase is heavily biased because of its uneven affinity towards the different template sequences. In addition, these techniques necessitate large amounts of RNA input.

This white paper discusses the straightforward, cost-effective, highly efficient “Capture and Amplification by Tailing and Switching” (CATS) to easily generate ready-to-sequence NGS libraries from picogram quantities of RNA.

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RNA-seqライブラリーキットの選択方法

Small RNAs

(Small non-coding RNA)

Input 10 pg-10 ng

Total RNA

(Whole transcriptome)

Input 100 pg-10 ng

mRNA

(Transcriptome)

Input 100 pg-10 ng

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

CATS RNA-seq Kit v2
(no polyA selection, no rRNA depletion modules)

CATS mRNA-seq Kit v2
(with polyA selection module)

CATS RNA-seq Kit v2
(no polyA selection, no rRNA depletion modules)

í注: 入力とは、単離されたsmall RNA、rRNA欠損またはポリ(A)選択されたRNA

Products

Cat. No.ProductFormatPrice
C05010043 CATS mRNA-seq Kit (with polyA selection) v2 x24
Diagenode’s CATS mRNA-seq Kit (with polyA selection) utilizes the innovative “Capture and Amplification by Tailing and Switching” (CATS), a...
24 rxns $1,395.00
C05010041 CATS RNA-seq Kit v2 x24
Diagenode’s CATS RNA-seq Kit utilizes the innovative “Capture and Amplification by Tailing and Switching” (CATS), a ligation-free method to...
24 rxns $1,360.00
C05010040 CATS Small RNA-seq Kit x24
Diagenode’s CATS small RNA-seq Kit utilizes the innovative “Capture and Amplification by Tailing and Switching” (CATS), a ligation-free met...
24 rxns $1,360.00
C05010042 CATS Total RNA-seq Kit (with rRNA depletion) v2 x24
Diagenode’s CATS total RNA-seq Kit (with rRNA depletion) utilizes the innovative “Capture and Amplification by Tailing and Switching” (CATS...
24 rxns $2,450.00

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