GSM5116019: Focused screen, upper bin, 400k cells; Homo sapiens; OTHER
Genomic DNA was prepared from sorted lower and upper bins and input samples containing 80,000 to 2 Mio cells per sample. Monarch genomic DNA purification kit (New England Biolabs) was used for gDNA preparations while including the RNase treatment. CROP/gRNA loci were amplified from gDNA using a nested PCR strategy with outer primers and barcoded inner primers containing Illumina adapters. Libraries were pooled and sequenced using an Illumina NextSeq 500 in high output mode with 8 reads to read out the i7 barcode, and 67 reads on Read1 to read through the stagger sequence and identify the gRNA. CRISPR guide RNA library sequencing