Improvement of bacterial transformation efficiency using plasmid artificial modification.
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.
Yasui
Kazumasa
K
The United Graduate School of Agricultural Science, Gifu University, Gifu, Japan.
Kano
Yasunobu
Y
Tanaka
Kaori
K
Watanabe
Kunitomo
K
Shimizu-Kadota
Mariko
M
Yoshikawa
Hirofumi
H
Suzuki
Tohru
T
D001644:Bifidobacterium / Q000235:genetics; D015254:DNA Modification Methylases / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D010957:Plasmids / Q000235:genetics*; D014169:Transformation, Bacterial
D015254:DNA Modification Methylases