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Fig. 4 | Clinical and Translational Medicine

Fig. 4

From: Single-cell sequencing and tumorigenesis: improved understanding of tumor evolution and metastasis

Fig. 4

Main approaches used for whole-transcriptome amplification of single cells. a The Tang method performs reverse transcription of mRNA for single-cell RNA-seq using an oligo-dT primer with an anchor sequence, then a poly-A tail is added to the 3′-end of the first cDNA and the second strand is synthesized using a different oligo-dT primer with a different anchor sequence; b Smart-seq and Smart-seq2 implement a template-switching step to increase the number of full-length cDNA transcripts with an intact 5′-end; c quartz-seq limits amplification of unwanted byproducts by removing excess primer with exonuclease I before second-strand synthesis and using suppression PCR to form hairpin structures that cannot be amplified; d cell expression by linear amplification and sequencing (CEL-Seq) includes a template-switching step and uses molecular barcodes and pooling of samples from multiple single cells prior to linear amplification; e single-cell tagged reverse transcription (STRT) permits multiplex sequencing of multiple cells in the same reaction using a template-switching mechanism to simultaneously introduce a molecular barcode and an upstream primer-binding sequence during reverse transcription; f quantitative single-cell RNA-seq generates full-length transcripts using template switching and incorporating random UMI (unique molecular identifier) sequences to label individual cDNA molecules and eliminate amplification bias [8]

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