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Nucleic Acids Res. 2009 Jan;37(1):e3. doi: 10.1093/nar/gkn884. Epub 2008 Nov 12.

Improvement of bacterial transformation efficiency using plasmid artificial modification.

Author information

1
The United Graduate School of Agricultural Science, Gifu University, Gifu, Japan.

Abstract

We have developed a method to improve the transformation efficiency in genome-sequenced bacteria, using 'Plasmid Artificial Modification' (PAM), using the host's own restriction system. In this method, a shuttle vector was pre-methylated in Escherichia coli cells, which carry all the putative genes encoding the DNA modification enzymes of the target microorganism, before electroporation was performed. In the case of Bifidobacterium adolescentis ATCC15703 and pKKT427 (3.9 kb E. coli-Bifidobacterium shuttle vector), introducing two Type II DNA methyltransferase genes lead to an enhancement in the transformation efficiency by five orders of magnitude. This concept was also applicable to a Type I restriction system. In the case of Lactococcus lactis IO-1, by using PAM with a putative Type I methyltransferase system, hsdMS1, the transformation efficiency was improved by a factor of seven over that without PAM.

PMID:
19004868
PMCID:
PMC2615632
DOI:
10.1093/nar/gkn884
[Indexed for MEDLINE]
Free PMC Article

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