scM&T-Seq

scM&T-Seq

Methylome and transcriptome sequencing from single-cells (scM&T-Seq) allows parallel analysis of both epigenetic and gene expression patterns from single-cells using Smart-Seq2 and scBS-Seq. scM&T-Seq is built upon G&T-Seq, but instead of using MDA for DNA sequencing, it uses scBS-Seq to determine DNA methylation patterns.

Single cells are isolated and individually lysed. The mRNAs are then captured with streptavidin-coupled mRNA capture primers to physically separate them from DNA strands. Smart-Seq2 uses reverse transcription with template switching and tagmentation to generate cDNA libraries from the mRNA. DNA libraries are prepared via scBS-Seq, which involves bisulfite conversion of DNA strands to identify methylated cytosines. Both libraries are now ready for sequencing.

Pros:

• Investigate links between epigenetic and transcriptional heterogeneity in single-cells
• Because DNA and RNA are physically separated and amplified independently, there is no need to mask coding sequences during analysis

Cons:

• Smart-Seq2 is not strand-specific and applicable to only poly(A)+ RNA
• Does not distinguish between 5mC and 5hmC

• scM&T-Seq: Angermueller C., Clark S. J., Lee H. J., Macaulay I. C., Teng M. J., et al. (2016) Parallel single-cell sequencing links transcriptional and epigenetic heterogeneity. Nat Methods 13: 229-232

1. Clark S. J., Lee H. J., Smallwood S. A., Kelsey G. and Reik W. (2016) Single-cell epigenomics: powerful new methods for understanding gene regulation and cell identity. Genome Biol 17: 72
2. Wen L. and Tang F. (2016) Single-cell sequencing in stem cell biology. Genome Biol 17: 71