Details

Autor(en) / Beteiligte

• Zhou, Yuexin
• Zhu, Shiyou
• Cai, Changzu
• Yuan, Pengfei
• Li, Chunmei
• Huang, Yanyi
• Wei, Wensheng

Titel

High-throughput screening of a CRISPR/Cas9 library for functional genomics in human cells

Ist Teil von

• Nature (London), 2014-05, Vol.509 (7501), p.487-491

Ort / Verlag

London: Nature Publishing Group UK

Erscheinungsjahr

2014

Quelle

EBSCOhost Psychology and Behavioral Sciences Collection

Beschreibungen/Notizen

• This study describes the construction of a focused CRISPR/Cas-based lentiviral library in human cells and a method of gene identification based on functional screening and high-throughput sequencing analysis. An effective knockout genomic screen The targeted modification of individual genes has become routine, with many alternative strategies available, but progress towards large-scale screening based on complete loss of gene expression has been slow. Arrayed screening based on subgenomic RNA (sgRNA) collections is a promising strategy, and here Yuexin Zhou et al . present a proof-of-concept demonstration of knockout screens in mammals using a focused CRISPR/Cas-based lentiviral sgRNA library in human cells and a method of gene identification based on functional screening and high-throughput sequencing analysis. Refinements of this technology could find broad application in the study of gene function and disease. Targeted genome editing technologies are powerful tools for studying biology and disease, and have a broad range of research applications 1 , 2 , 3 , 4 , 5 , 6 , 7 . In contrast to the rapid development of toolkits to manipulate individual genes, large-scale screening methods based on the complete loss of gene expression are only now beginning to be developed 8 , 9 . Here we report the development of a focused CRISPR/Cas-based (clustered regularly interspaced short palindromic repeats/CRISPR-associated) lentiviral library in human cells and a method of gene identification based on functional screening and high-throughput sequencing analysis. Using knockout library screens, we successfully identified the host genes essential for the intoxication of cells by anthrax and diphtheria toxins, which were confirmed by functional validation. The broad application of this powerful genetic screening strategy will not only facilitate the rapid identification of genes important for bacterial toxicity but will also enable the discovery of genes that participate in other biological processes.