Development of a genetically engineered Escherichia coli strain for plasmid transformation in Corynebacterium glutamicum
Development of a genetically engineered Escherichia coli strain for plasmid transformation in Corynebacterium glutamicum
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开发用于谷氨酸棒杆菌质粒转化的基因工程大肠杆菌菌株
DOI：
10.1016/j.mimet.2016.10.019
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发表时间：
2016-12-01
影响因子：
2.2
通讯作者：
Xu, Daqing
中科院分区：
文献类型：
--
作者：
Li, Hedan;Zhang, Lirong;Xu, Daqing
谷氨酸棒状杆菌的基因破坏和替换依赖于高的转化效率。cgllR-cglJR限制性体系是谷氨酸棒状杆菌细胞导入外源DNA的主要障碍。为了提高谷氨酸切菌的转化效率，将谷氨酸切菌ATCC 13032的cglIR-cglIIR-cglIM限制性修饰系统中编码甲基转移酶的cglIM基因进行染色体整合，并在大肠杆菌中表达，得到工程菌株E. coli AUl。大肠杆菌-大肠杆菌穿梭质粒pAU4电转化谷氨酸C ATCC 13032的实验表明，谷氨酸C对大肠杆菌TGl/pAU4中提取的pAU4 DNA的转化效率为1.80 +/- 0.21 × 10(2) cfu/ μ g质粒DNA的转化效率为5.22 +/- 033 × 10(6) cfu/ μ g质粒DNA的转化效率为5.22 +/- 033 × 10(6) cfu/ μ g。结果表明，大肠杆菌AU1能够将cglim特异性DNA甲基化模式赋予其驻留质粒，使质粒抵抗cglIR-cgIIIR限制并有效转移到C谷氨酸中。大肠杆菌AU1是一种有效转化谷氨酸C的中间宿主。(C) 2016年Elsevier B.V.出版
Gene disruption and replacement in Corynebacterium glutamicum is dependent upon a high transformation efficiency. The cgllR-cglJR restriction system is a major barrier to introduction of foreign DNA into Corynebacterium glutamicum cells. To improve the transformation efficiency of C. glutamicum, the cglIM gene encoding methyltransferase in the cglIR-cglIIR-cglIM restriction-modification system of C. glutamicum ATCC 13032 was chromosomally integrated and expressed in Escherichia coli, resulting in an engineered strain E. coli AUl. The electrotransformation experiments of C glutamicum ATCC 13032 with the E. coli-C glutamicum shuttle plasmid pAU4 showed that the transformation efficiency of C glutamicum with pAU4 DNA extracted from E. coli TGl/pAU4 was 1.80 +/- 0.21 x 10(2) cfu/mu g plasmid DNA, while using pAU4 DNA extracted from E. coli AUl/pAU4, the transformation efficiency reached up to 5.22 +/- 033 x 10(6) cfu/mu g plasmid DNA. The results demonstrated that E. coli AU1 is able to confer the cglIM-specific DNA methylation pattern to its resident plasmid, which makes the plasmid resistant to the cglIR-cgIIIR restriction and efficiently transferred into C glutamicum. E. coli AU1 is a useful intermediate host for efficient transformation of C glutamicum. (C) 2016 Published by Elsevier B.V.