Quartz-Seq

Quartz-Seq

The Quartz-Seq method optimizes whole-transcript amplification (WTA) of single cells. In this method, an RT primer with a T7 promoter and PCR target is first added to the extracted mRNA. RT synthesizes first-strand cDNA, after which the RT primer is digested by exonuclease I. Next, a poly(A) tail is added to the 3' ends of first-strand cDNA, along with a poly(dT) primer containing a PCR target. After second-strand generation, a blocking primer is added to ensure PCR enrichment in sufficient quantity for sequencing. Deep sequencing allows for accurate, high-resolution representation of the whole transcriptome of a single cell.

Similar methods: CEL-Seq, Drop-seq, MARS-Seq, CytoSeq, inDrop, Hi-SCL.

Pros:

• Single-tube reaction suitable for automation
• Digestion of RT primers by exonuclease I eliminates amplification of byproducts
• Short fragments and byproducts are suppressed during enrichment

Cons:

• PCR biases can underrepresent GC-rich templates
• Amplification errors caused by polymerases will be represented and sequenced incorrectly
• Targets smaller than 500 bp are preferentially amplified by polymerases during PCR

• Quartz-Seq: Sasagawa Y., Nikaido I., Hayashi T., Danno H., Uno K. D., et al. (2013) Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity. Genome Biol 14: R31

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