Discover-scTM WTA Kit V2 can use total RNA from 1-1000 cells or 10 pg- 10 ng total RNA as template, via first-strand cDNA synthesis and amplification, to achieve enough amount of sample for sequencing analysis, thereby solves the technical problems that sequencing analysis cannot be accomplished by regular mRNA-seq because of low RNA content in small samples such as single cell.
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Discover-scTM WTA Kit V2 is an updated version based on Discover-scTM WTA Kit. The detection sensitivity and volume compatibility have been significantly improved, and it is more suitable for detection of templates of low abundance and low concentration.
Low starting amount of template: Single cell or 10 pg total RNA can serve as starting templates for efficient amplification.
High sensitivity of amplification: Well optimized reaction system and LNA technology greatly increase the detection rate of low expression genes.
Good integrity of product: Full length cDNA is amplified by two end primers, to obtain information of full transcript, avoiding 5'and 3' preferences.
High success rate of operation: Treatment process of RNA sample is greatly reduced, to eliminate the risk of sample loss to the largest extent, and improve the success rate of operation.
Wide compatibility of volume: The compatible sample volume can be as large as 5 μl, compatible with different concentrations of template for amplification.
10 pg total RNA of 293T cells was amplified with Discover-scTM WTA Kit V2, and 1 μl cDNA amplification product was analyzed by Agilent 2100 Bioanalyzer. Oligo dT Primer was used as reverse transcription primers, and a linker sequence was added at the 3 'end of cDNA by Template-switching activity of the Discover-sc Reverse Transcriptase. Following PCR amplification employed the linker sequence, to obtain full-length cDNA amplification products, while avoiding the contamination of rRNA.
Vazyme-1 and Vazyme-2 refer to amplifications of single 293T cell by Discover-scTM WTA Kit V2; C-1 and C-2 refer to amplifications of single 293T cell by single cell transcriptome kit of C company. 1 ng cDNA amplification product were used to construct libraries using TruePrepTM DNA Library Prep Kit V2 for Illumina®. Following information are analyzations of these libraries.
The detection sensitivity of amplification products of Discover-scTM WTA Kit V2 was high, and the number of genes detected in single 293T cell was as high as 15000.
Left picture shows gene distribution results of the Vazyme-1 sequencing fragments, and right picture shows gene distribution results of the C-1 sequencing fragments. Data shows that amplification by Discover-scTM WTA V2 ensures good coverage of gene 5 'and 3' information.
Left picture shows correlation and repeatability analyzation of Vazyme-1 and Vazyme-2, and right picture shows correlation and repeatability analyzation of C-1 and C-2. The result data shows that the correlation and repeatability of our product is higher than that of C company.