Assembling the Streptococcus thermophilus clustered regularly interspaced short palindromic repeats (CRISPR) array for multiplex DNA targeting

Analytical Biochemistry
Lijun GuoZhiying Zhang

Abstract

In addition to the advantages of scalable, affordable, and easy to engineer, the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) technology is superior for multiplex targeting, which is laborious and inconvenient when achieved by cloning multiple gRNA expressing cassettes. Here, we report a simple CRISPR array assembling method which will facilitate multiplex targeting usage. First, the Streptococcus thermophilus CRISPR3/Cas locus was cloned. Second, different CRISPR arrays were assembled with different crRNA spacers. Transformation assays using different Escherichia coli strains demonstrated efficient plasmid DNA targeting, and we achieved targeting efficiency up to 95% with an assembled CRISPR array with three crRNA spacers.

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Citations

May 6, 2016·Frontiers in Plant Science·Zafar IqbalMuhammad Shafiq
Jul 3, 2015·Cell & Bioscience·Sean M RiordanShui Qing Ye
Sep 15, 2020·Molecular Biology Reports·Xin SongLian-Zhong Ai
Mar 9, 2018·Frontiers in Microbiology·Mengyuan HaoXiaojun Qu

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