Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Hiroshima, Japan.
FEMS Microbiol Lett. 2013 Jun;343(1):8-12. doi: 10.1111/1574-6968.12113. Epub 2013 Mar 15.
A transformation system for Moorella thermoacetica ATCC39073 was developed using thermostable kanamycin resistant gene (kanR) derived from the plasmid pJH1 that Streptococcus 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.
开发了一种用于热醋穆尔氏菌 ATCC39073 的转化系统，该系统使用源自粪肠球菌携带的质粒 pJH1 的耐热卡那霉素抗性基因 (kanR)。当带有其天然启动子的 kanR 与尿嘧啶营养缺陷型突变体一起引入到热醋穆尔氏菌 ATCC39073 中，并作为选择标记引入一个基因来补充尿嘧啶营养缺陷型时，由于 kanR 的转录水平低，它不能赋予卡那霉素抗性。然而，使用从热醋穆尔氏菌 ATCC39073 中克隆的甘油醛-3-磷酸脱氢酶启动子，显著提高了 kanR 的转录水平，导致细胞在超过 150μg/mL 的卡那霉素存在下生长。还证明了带有 G3PD 启动子的 kanR 可用于热醋穆尔氏菌 ATCC39073 野生型菌株的转化作为选择标记。
