GEM Generation and Barcoding v1

The Chromium Single Cell Gene Expression Solution upgrades short read sequencers to deliver a scalable microfluidic platform for 3’ digital gene expression by profiling 500-10,000 individual cells per sample. A pool of ~3,500,000 10x Barcodes are sampled separately to index each cell’s transcriptome. It is done by partitioning thousands of cells (or nuclei) into nanoliter-scale Gel Beads-in-emulsion (GEMs), where all generated cDNA share a common 10x Barcode. Dual Indexed libraries are generated and sequenced from the cDNA and 10x Barcodes are used to associate individual reads back to the individual partitions. This protocol outlines the process for generating Gel Beads-in-emulsion (GEMs) by combining barcoded Single Cell 3’ v3.1 Gel Beads, a Master Mix containing cells, and Partitioning Oil onto Chromium Next GEM Chip G. To achieve single cell resolution, cells are delivered at a limiting dilution, such that the majority (~90-99%) of generated GEMs contain no cell, while the remainder largely contain a single cell. This protocol details the GEM Generation and Barcoding procedures.