Open AccessEngineering of a functional thermostable kanamycin resistance marker for use in M oorella thermoacetica ATCC39073
Author(s) -
Iwasaki Yuki,
Kita Akihisa,
Sakai Shinsuke,
Takaoka Kazue,
Yano Shinichi,
Tajima Takahisa,
Kato Junichi,
Nishio Naomichi,
Murakami Katsuji,
Nakashimada Yutaka
Publication year - 2013
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/1574-6968.12113
Subject(s) - kanamycin , chemistry , microbiology and biotechnology , biology , genetics , bacteria
A transformation system for M oorella thermoacetica ATCC39073 was developed using thermostable kanamycin resistant gene ( kanR ) derived from the plasmid pJH1 that S treptococcus faecalis harbored. When kanR with its native promoter was introduced into uracil auxotrophic mutant of M . thermoacetica ATCC39073 together with a gene to complement the uracil auxotrophy as a selection marker, it did not give kanamycin resistance due to poor transcription level of kanR . However, the use of glyceraldehyde‐3‐phosphate dehydrogenase promoter cloned from M . thermoacetica ATCC39073 significantly improved transcription level of kanR and resulted in the cell growth in the presence of more than 150 μg mL −1 kanamycin. It was also demonstrated that kanR with G3PD promoter can be used as a selection marker for transformation of wild‐type strain of M . thermoacetica ATCC39073.