Efficient Multiplex Genome Editing in Streptomyces via Engineered CRISPR-Cas12a Systems

Frontiers in Bioengineering and Biotechnology
Jun ZhangYunzi Luo

Abstract

Streptomyces strains produce a great number of valuable natural products. With the development of genome sequencing, a vast number of biosynthetic gene clusters with high potential for use in the discovery of valuable clinical drugs have been revealed. Therefore, emerging needs for tools to manipulate these biosynthetic pathways are presented. Although the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas 9) system has exhibited great capabilities for gene editing in multiple Streptomyces strains, it has failed to work in some newly discovered strains and some important industrial strains. Additionally, the protospacer adjacent motif (PAM) recognition scope of this system sometimes limits its applications for generating precise site mutations and insertions. Here, we developed three efficient CRISPR-FnCas12a systems for multiplex genome editing in several Streptomyces strains. Each system exhibited advantages for different applications. The CRISPR-FnCas12a1 system was efficiently applied in the industrial strain Streptomyces hygroscopicus, in which SpCas9 does not work well. The CRISPR-FnCas12a2 system was used to delete large fragments ranging from 21.4 to 128 kb. Additionally, t...Continue Reading

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Citations

Mar 9, 2021·Frontiers in Bioengineering and Biotechnology·Zhenyu LiuYunzi Luo
Jul 16, 2021·Journal of Microbiology and Biotechnology·Ho Joung Lee, Sang Jun Lee

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Methods Mentioned

BETA
PCR
acetylation

Software Mentioned

Genewiz
BLAST

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