光合菌SDH2 hupT基因的突变与吸氢酶表达

利用三亲本杂交将自杀质粒ｐＳＥ８引入光合细菌Ｒｏｄｏｂａｃｔｅｒ ｓｐ．ＳＤＨ２０菌株，经过质粒上插入了ｋａｎ＾Ｒ基因的ｈｕｐＴ基因片段与受体基因组同源双交换，构建成ｈｕｐＴ插入突变株ＳＤＨＴ１和ＳＤＨＴ２。

Mutagenesis of hup T Gene and H 2 uptake Hydrogenase Expression in Photosynthetic Bacterium SDH2

HupT - insertion mutants, SDHT1 and SDHT2, were constructed by the homologous double exchange between the host, Rodobacter sp. SDH20, and the hup T gene fragment with a kan R gene inserted on pSE8, a suicide plasmid, which had been introduced into the host by using triparental conjugation. The HupT - mutants were verified by Southern hybridization. H 2 uptake hydrogenase activity analysis revealed that the H 2 uptake hydrogenase activities of HupT - mutants were two fold as that of wild type strain SDH20 when grown anaerobically in MN medium and aerobically in MG or MN medium, and had no distinct improvement relative to wild type strain when grown anaerobically in MG medium. The expression analysis of hup S confirmed that the HupT - mutants exhibited two to three fold increase of the expression of hup S relative to wild type strain when grown anaerobically in MN medium and aerobically in MG or MN medium, but no obvious increase when grown anaerobically in MG medium. These results demonstrate that hup T regulates negatively the synthesis of H 2 uptake hydrogenase in the photosynthetic bacterium strain SDH2.