Multiple genes deletion based on Cre-loxP marker-less gene deletion system for the strains from the genus of Pectobacterium.

Autor: Che S; Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China.; Plant Protection Institute, Jiangxi Academy of Agricultural Sciences, Nanchang, 330200, China., Zhuo Y; Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China., Yang L; Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China.; Institute of Grain Production, Yunnan Academy of Agricultural Science, Kunming, 650205, China., Wang H; Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China.; Institute of Agricultural Science of Suzhou, Taihu Lake District, Suzhou, 215155, China., Cui Z; Key Laboratory of Agricultural Environmental Microbiology, Ministry of Agriculture and Rural Affairs, College of Life Sciences of Nanjing Agricultural University, Nanjing, 210095, China., Fan J; Laboratory of Bacteriology, Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China. fanjq@njau.edu.cn.
Jazyk: angličtina
Zdroj: Biotechnology letters [Biotechnol Lett] 2024 Dec; Vol. 46 (6), pp. 1133-1142. Date of Electronic Publication: 2024 Jul 31.
DOI: 10.1007/s10529-024-03518-8
Abstrakt: Objective: To introduce the Cre-loxP system for constructing marker-less multiple-gene deletion mutants in Pectobacterium, overcoming limitations of antibiotic markers and enhancing the understanding of pathogenic mechanisms.
Results: Firstly, a plasmid named pEX18-Cre, containing a sacB sucrose suicide gene, was constructed to express Cre recombinase in Pectobacterium. Secondly, a mutant in which the loxP-Km fragment replaced the target gene was obtained through homologous recombination double-crossover with the chromosome. Finally, pEX18-Cre was introduced into the mutant to excise the DNA between the loxP sites, thereby removing the markers and achieving multiple gene deletions. By utilizing the Cre-loxP system, we successfully constructed multiple marker-less gene deletion mutants in Pectobacterium strains.
Conclusions: The Cre-loxP system efficiently creates marker-less multiple-gene deletion mutants, enhancing the study of Pectobacterium pathogenic mechanisms by overcoming antibiotic marker limitations.
(© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)
Databáze: MEDLINE
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