High throughput CRISPR screening in JURKAT cell line

We performed the survival-based and Notch signal-based genome-wide CRISPR screens in JURKAT cell lines. We established a JURKAT cell line that stably expressing HES1 promoter-driven EGFP as an indicator of Notch signaling. We carried out genome-wide CRISPR/Cas9-based screens in these cells to identify genes whose knockout decreases T-ALL cell proliferation and suppresses Notch signaling. For the survival screen, the infected cells were passaged every 3 days for 28 days, and samples of the initial and final cell populations were collected. For the Notch activity screen, the infected cells were sorted via fluorescence-activated cell sorting (FACS). The cell populations with GFP intensities in the lowest 20% (low Notch) and the highest 20% (high Notch) were collected and expanded, and the sorting process was repeated 3 times. The genomic DNA of the above cell populations was extracted and subjected to deep sequencing to evaluate the enrichment of genes in each population.