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SMART-Seq Stranded Kit

The SMART-Seq Stranded Kit is used to generate strand-specific RNA-seq libraries for Illumina® sequencing from 1–1,000 sorted cells or 10 pg–10 ng of purified total RNA. This kit incorporates Takara Bio's SMART (Switching Mechanism at the 5' end of RNA Template) technology and includes refinements to the SMARTer method for stranded RNA-seq that simplify the library preparation workflow and improve sequencing performance.

Brand: Takara

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Product CodeProduct Product Description Price (Excl gst) Qty Add
CLT634442 SMART-Seq® Stranded Kit, 12 Rxns SMART-Seq® Stranded Kit, 12 Rxns Login for price  
CLT634443 SMART-Seq® Stranded Kit, 48 Rxns SMART-Seq® Stranded Kit, 48 Rxns Login for price  
CLT634444 SMART-Seq® Stranded Kit, 96 Rxns SMART-Seq® Stranded Kit, 96 Rxns Login for price  

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The SMART-Seq Stranded Kit is used to generate strand-specific RNA-seq libraries for Illumina® sequencing from 1–1,000 sorted cells or 10 pg–10 ng of purified total RNA. This kit incorporates Takara Bio's SMART (Switching Mechanism at the 5' end of RNA Template) technology and includes refinements to the SMARTer method for stranded RNA-seq that simplify the library preparation workflow and improve sequencing performance. In addition, contrary to the SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing and SMART-Seq HT Kit, reverse transcription is initiated using random priming instead of oligo(dT) priming, thus capturing the full transcriptome instead of only the polyadenylated fraction. The SMART-Seq Stranded Kit was specifically designed to deliver highly sensitive and reproducible data from single cells while keeping the workflow short and user friendly. The kit does not require additional rRNA removal methods or kits and produces sequencing libraries that retain strand-of-origin information. The integrated removal of cDNAs derived from rRNA-typically present in high abundance following cDNA synthesis from total RNA inputs-makes the workflow extremely sensitive, yielding data that is highly reproducible with low mapping to rRNA.

 

Overview

  • Simple workflow to generate stranded Illumina sequencing-ready libraries in 7 hours
  • Appropriate for analysis of 1–1,000 intact mammalian cells or 10 pg–10 ng of low- or high-quality mammalian total RNA 
  • Reproducible, accurate detection of full-length coding and noncoding transcripts from single cells or total RNA 
  • Comparable sensitivity to our gold-standard SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing.

Interested in more data and FAQs about this product? Visit the NGS Learning Center.

Applications

  • Generation of Illumina sequencing-ready libraries in 7 hours from small quantities of mammalian cells (1–1,000 cells) or total RNA (10 pg–10 ng)
  • Whole transcriptome sequencing with strand-of-origin information

 

SMART-Seq Stranded Kit workflow
 

Schematic of workflow for the SMART-seq Stranded Kit

Figure 1. Schematic of the SMART-Seq Stranded Kit workflow.

 

SMART-Seq Stranded Kit performs well across a wide range of inputs

Sequencing alignment metrics for A375 total RNA and cells
Input Total RNA 1,000 cells 500 cells 100 cells 10 cells 5 cells 1 cell
Number of reads (pairs) 6,000,000 6,000,000 6,000,000 6,000,000 6,000,000 6,000,000 5,873,974
Number of transcripts >1 FPKM 13,260 13,294 13,583 13,520 12,726 12,602 11,540
Number of transcripts >0.1 FPKM 21,334 21,113 21,365 21,145 20,550 18,888 15,815
Proportion of reads (%):
Exonic 34.7 36.4 39.2 42.7 36.7 36.2 37.3
Intronic 29.6 29.3 27.7 28.3 34.0 30.4 21.1
Intergenic 14.2 13.4 12.2 12.9 16.7 16.8 10.1
rRNA 7.0 11.4 11.5 6.3 3.6 4.9 7.1
Mitochondrial 4.1 3.5 3.7 4.9 3.8 4.4 4.6
Overall mapping (%) 89.6 93.9 94.3 95.1 94.9 92.7 80.2
Duplicate rate (%) 37.3 45.2 40.3 46.1 52.5 72.2 78.5
lncRNA mapping:
Number of mapped reads (%) 7.2 10.4 10.8 9.4 8.7 8.6 7.3
lncRNA transcripts detected 5,395 4,687 4,565 5,439 5,440 4,983 2,802

Figure 2: High reproducibility across cell input amounts. A375 cells isolated by FACS were used to generate RNA-seq libraries with the SMART-Seq Stranded Kit. Input varied from 1 cell to 1,000 cells, with two replicates per input of 5–1,000 cells and 12 replicates for the single cells. For comparison, two aliquots of 1,000 cells were used for total RNA purification and then used for library preparation.

 

SMART-Seq Stranded Kit matches the sensitivity of SMART-Seq v4 technology
 

Similar sensitivity and reproducibility between SMART-Seq v4 and SMART-Seq Stranded kits

Figure 3. Comparison between SMART-Seq v4 and SMART-Seq Stranded kits. Single cells (K562) isolated by FACS were used to generate RNA-seq libraries with the SMART-Seq Stranded Kit (Stranded) and a SMART-Seq v4 Kit (SSv4; cDNA from this kit was further processed with a Nextera® XT DNA Library Preparation Kit). Panel A. The overlap in the total number of transcripts identified (FPKM >1) by each kit was analyzed and shown to be 83.8%. Panel B. The number of transcripts identified (FPKM >1) in individual cells was similar between the two kits, with a tighter range across cells processed with the SSv4 kit. Panel C. The reproducibility (Pearson correlation) of transcript expression levels across all cells from each kit was similar, although slightly higher and more consistent across cells processed with the Stranded kit.

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