An optimization system for isolating and sequencing of single human colon cancer cells

Single cell sequencing (SCS) is a promising approach for precisely digging into the genetic heterogeneity at single cells level. Single cell whole genome/exome sequencing (scWGS/scWES) and single cell whole transcriptome sequencing (scRNA-seq) are methods of SCS and have been applied in various studies of solid tumors, e.g. liver cancer, breast cancer. However, no scRNA-seq-related studies of human colon-cancer tissue-samples have been conducted. In this study, we developed a modified and efficient colon-cancer-tissue SCSs system (named CCTSs-SCSs system) by combining four optimization technologies: tissue digestion, live and intact single-cells capture, single-cell RNA or genome amplification and sequencing, and bioinformatics analysis. This advanced system is applicable to both scRNA-seq and scWGS/scWES. Using this system, we successfully completed several single cells scRNA-seq and scWES, and found three key points of this system: an improved digestion system, FACS-based dead-cell-removal method for scRNA-seq, 20 cycles for scRNA amplification. Our CCTSs-SCSs system provides a reliable and efficient method of scRNA-seq or scWGS/scWES for human colon cancer tissue and contributes to single-cell level and precise studies for colon cancer. 5 cells for beads-based DCR kit, and 5 cells for FACS-based DCR kit