Pre-existing RNA and ATAC modalities of single-cell tri-modal CRISPR screen of Homo sapiens: A375 melanoma Vemurafenib resistance
A375-dCas9-BFP-KRAB cells were transduced with a sgRNA library at a low MOI and BFP+ GFP+ double positive cells were sorted out. Cells were cultured for additional 6 days with either DMSO or 1uM Vemurafenib treatment. Fixed cells underwent Tn5 tagmentation in bulk, reverse transcription in bulk, two rounds of indexing ligation, and finally indexing PCR. SgRNA libraries were further separated and enriched by another round of PCR. A step-by-step experimental protocol of PerturbFate library preparation is included in the supplementary material of the publication. Nascent RNA part of the Tri-omics library have unique i7 barcodes, while pre-existing RNA and ATAC parts of the same single cell share the same cell barcode, and need computational demultiplexing of reads of each modality in the first step.