Hi-SCL

Hi-SCL

High-throughput single-cell labeling (Hi-SCL) generates transcriptome profiles for thousands of single-cells using custom microfluidics system similar to Drop-Seq and inDrops. Single cells from cell suspension are isolated into droplets containing lysis buffer. After cell lysis, cell-droplets are then fused with a droplet containing cell-specific barcodes and another droplet with enzymes for reverse transcription. Droplets from all the wells are pooled and subjected to isothermal reactions for reverse transcription. The barcode-oligos anneal to poly(A) mRNAs and act as primers for reverse transcriptase. Now that each mRNA strand have cell-specific barcodes, droplets are broken and mRNAs purified. The 3' ends of the cDNA strands are ligated with adaptors, amplified, annealed with indexed primers, and amplified further before sequencing.

Pros:

• High throughput single-cell transcriptome profiling using microfluidics system
• Low cost - $0.1 per cell (experiment with 100 cells)
• Highly scalable to larger cell quantities
• No fragmentation step

Cons:

• Lack of Unique Molecular Identifier (UMI) in oligos may create amplification noise
• Droplets may contain two cells or two different types of barcodes

• Hi-SCL: Rotem A., Ram O., Shoresh N., Sperling R. A., Schnall-Levin M., et al. (2015) High-Throughput Single-Cell Labeling (Hi-SCL) for RNA-Seq Using Drop-Based Microfluidics. PLoS One 10: e0116328

1. Mato Prado M., Frampton A. E., Stebbing J. and Krell J. (2016) Single-cell sequencing in cancer research. Expert Rev Mol Diagn 16: 1-5
2. Rotem A., Ram O., Shoresh N., Sperling R. A., Goren A., et al. (2015) Single-cell ChIP-seq reveals cell subpopulations defined by chromatin state. Nat Biotechnol