STRT

STRT

STRT-Seq is a method similar to CEL-Seq that involves unique barcoding and sample pooling to overcome the challenges of samples with limited material. , In this method, single cells are first picked in individual tubes, where first-strand cDNA synthesis occurs using an oligo(dT) primer with the addition of 3–6 cytosines. A helper oligonucleotide promotes template switching, which introduces the barcode into the cDNA. The barcoded cDNA is amplified by single-primer PCR. Deep sequencing allows for accurate transcriptome determination of individual cells.

Pros:

• Barcoding and pooling allows for multiplexing and studying many different single cells at a time
• Sample handling and the potential for cross-contamination are greatly reduced due to using a single tube per cell

Cons:

• PCR biases can underrepresent GC-rich templates
• Nonlinear PCR amplification can lead to biases affecting reproducibility
• Amplification errors caused by polymerases will be represented and sequenced incorrectly
• Loss of accuracy due to PCR bias
• Targets smaller than 500 bp are amplified preferentially by polymerases during PCR

• STRT: Islam S., Kjallquist U., Moliner A., Zajac P., Fan J. B., et al. (2011) Characterization of the single-cell transcriptional landscape by highly multiplex RNA-seq. Genome Res 21: 1160-1167

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