Dataset Information

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

ABSTRACT: The 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.

SUBMITTER: Li XT

PROVIDER: S-EPMC5171832 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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tCRISPRi: tunable and reversible, one-step control of gene expression.

Li Xin-Tian XT Jun Yonggun Y Erickstad Michael J MJ Brown Steven D SD Parks Adam A Court Donald L DL Jun Suckjoon S

Scientific reports 20161220

The 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 syst ...[more]

PMID: 27996021

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Dataset Information

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

Publications

tCRISPRi: tunable and reversible, one-step control of gene expression.

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