1. High-performance CRISPR-Cas12a genome editing for combinatorial genetic screening.
- Author
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Gier RA, Budinich KA, Evitt NH, Cao Z, Freilich ES, Chen Q, Qi J, Lan Y, Kohli RM, and Shi J
- Subjects
- Animals, Cell Proliferation, Epigenesis, Genetic, Gene Library, Genetic Engineering, Genome, Human, HEK293 Cells, Humans, K562 Cells, Mice, NIH 3T3 Cells, Protein Domains, RNA, Guide, CRISPR-Cas Systems genetics, Bacterial Proteins genetics, CRISPR-Associated Proteins genetics, CRISPR-Cas Systems, Endodeoxyribonucleases genetics, Gene Editing, Gene Expression Regulation, Leukemic, Leukemia genetics
- Abstract
CRISPR-based genetic screening has revolutionized cancer drug target discovery, yet reliable, multiplex gene editing to reveal synergies between gene targets remains a major challenge. Here, we present a simple and robust CRISPR-Cas12a-based approach for combinatorial genetic screening in cancer cells. By engineering the CRISPR-AsCas12a system with key modifications to the Cas protein and its CRISPR RNA (crRNA), we can achieve high efficiency combinatorial genetic screening. We demonstrate the performance of our optimized AsCas12a (opAsCas12a) through double knockout screening against epigenetic regulators. This screen reveals synthetic sick interactions between Brd9&Jmjd6, Kat6a&Jmjd6, and Brpf1&Jmjd6 in leukemia cells.
- Published
- 2020
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