Create a workflow for CRISPR genome-wide knockout screen: (1) GeCKO/Brunello sgRNA library design (4-6 sgRNAs per gene), (2) lentiviral packaging in HEK293T, (3) target cell transduction at MOI~0.3, (4) puromycin selection for integration, (5) apply selective pressure (drug treatment vs vehicle control), (6) harvest surviving cells, (7) genomic DNA extraction, (8) PCR amplification of sgRNA cassette, (9) NGS sequencing, (10) MAGeCK/BAGEL2 analysis for gene-level enrichment/depletion scoring. Include representation at each step.
