Developing a broadly applicable pyrF-based genetic manipulation system in Acinetobacter baumannii
Developing a broadly applicable pyrF-based genetic manipulation system in Acinetobacter baumannii
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DOI：
10.1101/2020.11.05.370742
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Rui Wang
中科院分区：
文献类型：
--
作者：
Run Xu;Can Gao;Shuqi Wu;Mengjiao Su;Chengfu Su;Xu Jia;Rui Wang
Acinetobacter baumannii is an emergency pathogenic bacterium for its multidrug-resistance and high mortality rates after infection. In-depth genetic analysis of A. baumannii virulence and drug-resistant genes is highly desirable. Existing methods for genetic manipulation of A. baumannii mainly rely on the use of antibiotic as the selectable marker, and the sacB/sucrose as the counter-selectable marker, which is inconvenient and inappropriate for all research of A. baumannii. Based on the highly conserved pyrF gene and its conserved 500bp-flanking sequence, we quickly and easily generated the pyrF-deleted mutants as the uracil auxotrophic host strain in three model strains and 11 clinical strains. The pyrF-carried vectors constructed for gene editing with pyrF/5-FOA as the counter-selection were conveniently and time-saving in these pyrF-deleted mutants. Utilizing the pyrF-based genetic manipulation system, we easily and efficiently modified the cas gene and CRISPR sequence of I-F CRISPR-Cas system in A. baumannii AYE, and detected the CRISPR interference and adaptation in these mutants. In summary, the pyrF-based genetic manipulation system could be broadly applicable used for efficiently maker-less gene editing in most A. baumannii strains.
