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Ultra low input mRNA-Seq Kit

제조사	제품코드	제품명	용량	가격 (부가세별도)	비고
Clontech	634888	SMART-Seq® v4 Ultra® Low Input RNA Kit for Sequencing	12회 (Package)	가격문의	재고 소진 후 종매
Clontech	634889	SMART-Seq® v4 Ultra® Low Input RNA Kit for Sequencing	24회 (Package)	가격문의	재고 소진 후 종매
Clontech	634890	SMART-Seq® v4 Ultra® Low Input RNA Kit for Sequencing	48회 (Package)	가격문의	재고 소진 후 종매

Product index

PGT-A
- Embgenix™ PGT-A Kit (..
SMARTer RNA-Seq 개요
mRNA-Seq (Single cell)
Stranded RNA-seq
Small RNA-Seq
- Small RNA-Seq Kit
Immune-Seq
기타 RNA-Seq
- Full length cDNA 합성
- Target RNA Capture for..
DNA-seq 개요
- [기술개요] ThruPLEX & ..
- DNA-Seq 선택가이드
ChIP-Seq / Meth-Seq
Low Input DNA-Seq
High input DNA-seq
- ThruPLEX® DNA-Seq HV
- ThruPLEX® Tag-Seq HV
Single cell DNA-Seq
- PicoPLEX® Single Cell..
Whole Genome Amplification..
Index Kit
NGS Library 정량
- Library Quantification..
NGS Sample preparation
NGS 분석용 PCR 효소
NGS 관련 장비
[Legacy] NGS Products
- (Legacy) Human TCR a/b..
- (Legacy) PicoPLEX® WG..

* SMART-Seq^® v4 Ultra^® Low Input RNA Kit for Sequencing 제품과 SMART-Seq^® v4 PLUS Kit 제품은 각각 SMART-Seq^® mRNA, SMART-Seq^® mRNA LP로 리뉴얼 되었습니다.
* UMIs 부가로 정확도와 신뢰도를 높인 신제품, SMART-Seq^® mRNA LP (with UMIs)가 출시되었습니다.

* PLUS kit는 샘플로부터 lllumina^® 분석용 library 제작을 위한, library preparation kit와 single-use unique dual index (UDI)를 포함하고 있습니다.

* 자동화 시스템 (Apollo™ Library Prep System (시스템은 판매종료))에 적용가능한 시약
- SMART-Seq^® v4 Reagent Kit for the SMARTer™ Apollo™ System (Code 640170)

[관련 정보자료]
- SMART-Seq v4 Single cell RNA-seq 온라인 세미나 바로보기
- SMART-Seq v4 Ultra Low Input Kit를 이용한 Single cell RNA-Seq 논문 리스트 보기

10 pg 수준의 극소량 total RNA에도 적용 가능
Unparalleled sensitivity and reproducibility
Single-tube workflows - cDNA 합성부터 library 제작 과정을 각각 single tube에서 진행할 수 있어, 샘플 손실이나 오염 등 실험 과정에서 발생할 수 있는 오류 최소화
High-quality RNA-seq data - total RNA로부터 가장 많은 수의 유전자 분석
Increase your sequencing power - 많은 샘플을 분석할 수 있는 Unique dual index (UDI) 사용
High library yield - 높은 yield의 library를 제작할 수 있어, NovaSeq™ system과 같은 high-throughput sequencer에 적용하거나, 여러 차례 분석 가능

제품설명

SMART-Seq^® v4 Ultra^® Low Input RNA Kit for Sequencing (SSv4)와 SMART-Seq^® v4 PLUS kit (SSv4 PLUS)는 극소량 (Ultra-Low input) 샘플을 위한 SMARTer Ultra Low kit 시리즈의 4세대 제품으로, 극소량 샘플을 가장 민감도 있게 분석할 수 있다. SSv4 PLUS 제품을 이용하면, 1,000개 이하의 intact한 세포 혹은 10 pg - 10 ng의 high quality RNA (RIN > 8)로부터 oligo(dT) primer를 이용해 고품질 full-length cDNA를 제조한 후, lllumina^® 분석용 library 제작까지 진행할 수 있다.
본 제품은 SMART (Switching Mechanism at 5' End of RNA Template) 기술에 Locked Nucleic Acid (LNA) 기술을 접목함으로써 template switching 효율을 더욱 향상시켰다. 따라서, 이전 세대의 제품보다 더욱 많은 수의 유전자를 검출할 수 있어 전반적인 전사체 연구 (full transcriptome analysis)에 있어 매우 유용하다. SSv4는 높은 재현성과 gene-body coverage 뿐 아니라 GC-rich 전사체도 정확하게 cDNA로 합성할 수 있다.
SMART-Seq^® v4 PLUS kit (SSv4 PLUS)에 포함되어 있는 library preparation kit는 enzymatic fragmentation와 stem-loop adaptor를 함께 사용하여, 2시간 내 single tube에서 purification 과정 없이 cDNA로부터 library를 제작할 수 있다. 따라서 샘플의 손실이나 오염 등 실험 과정에서 발생할 수 있는 오류를 최소화 할 수 있다. 이 과정을 통해 제작된 library는 SMART-Seq2 등 기존의 방법 및 당사 기존 제품 (SMART-Seq^® v3)에 비해 재현성이 높고, 훨씬 우수할 뿐 아니라, 높은 yield로 제작되어 high-throughput sequencer에 적용하거나, 여러 차례 분석할 수 있다.

그림 1. SMART기술을 이용한 cDNA 합성 과정

The SMART-Seq^® v4 Oligonucleotide, 3’ SMART-Seq^® CDS Primer II A, and PCR Primer II A all contain a stretch of identical sequence. The black star indicates a chemical block on the 5’ end of the oligonucleotide.

그림 2. SSv4를 이용했을 때 높은 민감도와 mapping 효율

Replicate libraries were generated from 10 pg Mouse Brain Total RNA using the SMART-Seq^® v4 kit, the SMARTer^® Ultra Low v3 kit, or the SMART-Seq2 method. 18 PCR cycles were used to amplify cDNA libraries with the SMART-Seq2 method and SMARTer^® Ultra Low v3 kit; however, only 17 PCR cycles were needed for the SMART-Seq^® v4 libraries.

그림 3. 기존 방법인 SMART-Seq2, SSv3와 비교했을 때, SSv4의 높은 gene coverage

Gene body coverage shown is the average of two replicate libraries prepared from 10 pg Mouse Brain Total RNA using the three different cDNA synthesis methods. The SMARTer^® Ultra Low v3 kit produced a slight 3′ bias, and the SMART-Seq^® v4 kit produced a slight 5′ bias; however, the overall coverage was fairly even.

그림 4. 낮은 input의 샘플 사용에도 높은 재현성

FPKMs from replicate libraries generated from 10 pg of Mouse Brain Total RNA using the SMART-Seq^® v4 kit, the SMARTer^® Ultra Low v3 kit method were compared. For transcripts with FPKM <100, the correlation between replicates was much higher for the SMART-Seq^® v4 kit (Pearson R = 0.739; Panel B) compared to the SMARTer Ultra Low v3 (Pearson R = 0.376; Panel A). For all transcripts (shown in the scatterplots on the right) the correlation between replicates was high for each of the three methods (Pearson R between 0.911-0.972), though the SMART-Seq^® v4 kit did have the highest correlation. Transcripts represented in only one replicate can be seen along the X- and Y-axes of the scatter plots showing all transcripts.

그림 5.SSv4-HT PLUS를 이용한 library에서 Nextera^® XT보다 더 높은 yield와 유전자 검출

(Panel A) SSv4 chemistry was used to produce cDNA, in triplicate, from 10 pg of Mouse Brain Total RNA. Illumina-compatible libraries were then generated using the SMART-Seq^® Library Prep Kit (not sold separately) portion of the SSv4 PLUS kit or Nextera^® XT and sequenced on a NextSeq 500. The reads were normalized to 4M paired-end reads and analyzed as described in the techical note.
(Panel B) The library yield obtained with the SSv4 PLUS kit is higher than the yield obtained when using Nextera^® XT.
(Panel C) The number of genes identified was also higher for the SSv4 PLUS kit compared to Nextera^® XT.

적용

극소량 샘플로부터 full-length transcriptome 분석 (transcript isoforms, gene fusions, point mutations 등)을 위해, cDNA 합성 과정에서 사용할 수 있다. 본 제품을 이용해 합성한 cDNA는 SMART-Seq Library Prep kit (SSv4 PLUS kit 내 포함, 별도 구매 불가) 를 이용하여 Illumina^® 분석용 sequencing library로 제작할 수 있다. 혹은 Ion Xpress Plus Fragment Library Kit를 이용하여, Ion Torrent sequencing를 진행할 수 있다.

자세한 구성은 CoA를 참고하세요.

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Keyword : Single cell,단일세포,싱글셀,single cell analysis,single cell NGS,full length,Ultra low input,low input,극소량,mRNA,mRNA-seq,cDNA,cDNA-seq,NGS,Next Generation Sequencing,SMART,smart-seq,SSv4

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