GSM6529423: 50gLib_TeloHAEC_150K_Dialout_1; Homo sapiens; OTHER
Cells were either loaded at the maximum recommended concentration (16,000 cells/lane, "16k"), or "superloaded" at 150,000 cells per 10X Chromium controller lane ("150k"). 1 lane was used for each cell line/loading condition for the 50gene library, and 2 per cell line (superloading only) for the 200 gene library. scRNA-seq and dialout libraries were prepared as described below. 1) scRNA-seq libraries were prepared using 3' scRNA-seq V3 kits (10X Genomics), according to manufacturer instructions. For the Eahy926 50 gene 150k loading, three techical replicate libraries were made from the amplified cDNA from a single lane (denoted "scRNAseq.1, .2 or .3, in library names & "_A2", "_A3" & "_A4" in fastq filenames). Note that the 10x Indexes were a mix of 4 oligos for each library (each starting with a different base to avoid sequencing biases). Thus each scRNAseq library has 4 fastq files for each read direction (denoted _1, _2, _3 & _4 in fastq file names). 2) From the initial amplified cDNA, we used a two stage PCR protocol to generate “dialout” libraries, for each lane. Because the CROP-seq vector expresses a PolII polyadenylated transcript that ends just downstream of the guide sequence, the dialout libraries identify the sgRNA sequences associated with each droplet.