Illumina paired end sequencing
Human gastric cancer tissue
Single cell encapsulation and cDNA libraries were prepared by Chromium™ Single Cell 5' Reagent Kits and Chromium™ Single Cell A Chip Kit version 1. Cells in suspension are counted and loaded into individual wells of 10X Chromium Single Cell chip. Single cells are then encapsulated into Gel in-beads emulsions (GEM) by 10X Chromium Controller. Single Cell 5' Reagent Kits is used to perform downstream steps, in brief as follows: reverse transcription is performed on GEMs, followed by cDNA clean up and amplification. The double-stranded cDNA will then go through enzymatic fragmentation, adapter ligation, index PCR and SPRIselect size selection per manufacturer's protocol. Library size and concentration is determined by Qubit, quantitative PCR and Bioanalyzer assays. The libraries are denatured and diluted to optimal concentration. Illumina NovaSeq reagent kit is used for sequencing run at 150 bp for read 1 and read 2, each 10X sample index set comprises of 4 indexes. Sequencing was performed by Centre for PanorOmic Sciences, The University of Hong Kong using Novaseq 6000.