Development of a fast and easy method for Escherichia coli genome editing with CRISPR/Cas9.

Abstract:

BACKGROUND:Microbial genome editing is a powerful tool to modify chromosome in way of deletion, insertion or replacement, which is one of the most important techniques in metabolic engineering research. The emergence of CRISPR/Cas9 technique inspires various genomic editing methods. RESULTS:In this research, the goal of development of a fast and easy method for Escherichia coli genome editing with high efficiency is pursued. For this purpose, we designed modular plasmid assembly strategy, compared effects of different length of homologous arms for recombination, and tested different sets of recombinases. The final technique we developed only requires one plasmid construction and one transformation of practice to edit a genomic locus with 3 days and minimal lab work. In addition, the single temperature sensitive plasmid is easy to eliminate for another round of editing. Especially, process of the modularized editing plasmid construction only takes 4 h. CONCLUSION:In this study, we developed a fast and easy genome editing procedure based on CRISPR/Cas9 system that only required the work of one plasmid construction and one transformation, which allowed modification of a chromosome locus within 3 days and could be performed continuously for multiple loci.

journal_name

Microb Cell Fact

journal_title

Microbial cell factories

authors

Zhao D,Yuan S,Xiong B,Sun H,Ye L,Li J,Zhang X,Bi C

doi

10.1186/s12934-016-0605-5

subject

Has Abstract

pub_date

2016-12-01 00:00:00

pages

205

issue

1

issn

1475-2859

pii

10.1186/s12934-016-0605-5

journal_volume

15

pub_type

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