Genome-Wide CRISPR/Cas9 Screening for High-Throughput Functional Genomics in Human Cells
It is highly desirable to identify gene’s function in a high-throughput fashion, and the CRISPR/Cas9 system has been harnessed to meet such a need. Here, we describe a general method to generate genome-scale lentiviral single-guide RNA (sgRNA) library and conduct a pooled function-based screening in human cells. This protocol would be of interest to researchers to rapidly identify genes in a variety of biological processes.
Key wordsCRISPR-Cas9 system High-throughput Knockout Screening sgRNA
We thank J. Xi for providing the Cas9-encoding construct, and D. Trono for aid with the lentivirus packaging system. We thank Peking University High-throughput Sequencing Center operated by BIOPIC. This work was supported by funds from the National Basic Research Program of China (2010CB911800), the National Science Foundation of China (NSFC31170126, NSFC31070115), and the Peking-Tsinghua Centre for Life Sciences. Shiyou Zhu and Yuexin Zhou contributed equally to this work.