title: Development of a genetically engineered Escherichia coli strain for plasmid transformation in Corynebacterium glutamicum.
abstract: Gene disruption and replacement in Corynebacterium glutamicum is dependent upon a high transformation efficiency. The cglIR-cgIIR 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 AU1. The electro-transformation 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 TG1/pAU4 was 1.80±0.21×102cfu/μg plasmid DNA, while using pAU4 DNA extracted from E. coli AU1/pAU4, the transformation efficiency reached up to 5.22±0.33×106cfu/μg 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-cglIIR restriction and efficiently transferred into C. glutamicum. E. coli AU1 is a useful intermediate host for efficient transformation of C. glutamicum.
authors: College of Life Sciences, Agricultural University of Hebei, Baoding 071001, China.
authors: Hedan
authors: H
authors: Li
authors: Biological Control Centre of Plant Diseases and Pests of Hebei, Agricultural University of Hebei, Baoding 071001, China.
authors: Lirong
authors: L
authors: Zhang
authors: Biological Control Centre of Plant Diseases and Pests of Hebei, Agricultural University of Hebei, Baoding 071001, China; Plant Science and Technology College, Beijing University of Agriculture, Beijing 102206, China. Electronic address: 1787421502@qq.com.
authors: Wei
authors: W
authors: Guo
authors: College of Life Sciences, Agricultural University of Hebei, Baoding 071001, China. Electronic address: daqingxu2013@126.com.
authors: Daqing
authors: D
authors: Xu
mesh_terms: D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D048230:Corynebacterium glutamicum / Q000201:enzymology / Q000235:genetics*; D019175:DNA Methylation; D015280:DNA Restriction-Modification Enzymes; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D008780:Methyltransferases / Q000235:genetics; D010957:Plasmids / Q000235:genetics*; D014169:Transformation, Bacterial
chemical_list: D015280:DNA Restriction-Modification Enzymes; D004269:DNA, Bacterial; D008780:Methyltransferases
keywords: Corynebacterium glutamicum; Intermediate host; Plasmid transformation; cglIM-specific DNA methylation