TCRISPRi: Tunable and reversible, one-step control of gene expression

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题名:	TCRISPRi: Tunable and reversible, one-step control of gene expression
作者:	田溶根;Li, Xin-tian;Jun, Yonggun;Erickstad, Michael J.;Brown, Steven D.;Parks, Adam;Court, Donald L.;Jun, Suckjoon
贡献者:	理學院物理學系
关键词:	14;14/19;14/35;38;38/35;42/35;631/1647/1511;631/326/41/2173;639/624/1020;96/63;Arabinose;Arabinose - metabolism;Arabinose operon;Bacterial Proteins - genetics;Cellular apoptosis susceptibility protein;CRISPR;CRISPR-Cas Systems;Gene deletion;Gene Expression;Gene Expression Regulation, Bacterial;Gene Knockout Techniques;Genetic Engineering - methods;Humanities and Social Sciences;multidisciplinary;Operon;Promoter Regions, Genetic;Quantitative research;Science;Titration
日期:	2016-12-20
上传时间:	2026-04-23 14:39:14 (UTC+8)
出版者:	Nature Publishing Group;London: Springer Science and Business Media LLC
摘要:	摘要： AbstractThe ability to control the level of gene expression is a major quest in biology. A widely used approach employs deletion of a nonessential gene of interest (knockout), or multi-step recombineering to move a gene of interest under a repressible promoter (knockdown). However, these genetic methods are laborious, and limited for quantitative study. Here, we report a tunable CRISPR-cas system, “tCRISPRi”, for precise and continuous titration of gene expression by more than 30-fold. Our tCRISPRi system employs various previous advancements into a single strain: (1) We constructed a new strain containing a tunable arabinose operon promoter PBAD to quantitatively control the expression of CRISPR-(d)Cas protein over two orders of magnitude in a plasmid-free system. (2) tCRISPRi is reversible, and gene expression is repressed under knockdown conditions. (3) tCRISPRi shows significantly less than 10% leaky expression. (4) Most important from a practical perspective, construction of tCRISPRi to target a new gene requires only one-step of oligo recombineering. Our results show that tCRISPRi, in combination with recombineering, provides a simple and easy-to-implement tool for gene expression control, and is ideally suited for construction of both individual strains and high-throughput tunable knockdown libraries. 其他題名： Sci Rep 出版者： London: Springer Science and Business Media LLC 出版日期： 2016-12-20 出處： Scientific Reports, 2016-12, Vol.6 (1), p.39076--39076, Article 39076 資源來源： Publicly Available Content Database 版權： The Author(s) 2016 版權： Copyright Nature Publishing Group Dec 2016 版權： Copyright © 2016, The Author(s) 2016 The Author(s) 識別號： ISSN: 2045-2322 識別號： EISSN: 2045-2322 識別號： DOI: 10.1038/srep39076 識別號： PMID: 27996021
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