转座子Tn5对葡萄根癌病生防菌MI15的诱变

Transposon Mutagenesis of Anti-grapevine Agrobacterium Radiobactor MI15

以含自杀性质粒pMO75::Tn5的铜绿假单胞菌PAO1826和含自杀性质粒pSUP2021::Tn5的大肠杆菌S17-1为供体菌,以葡萄根癌病生防菌MI15为受体菌,进行接合杂交.当供体菌和受体菌菌体数量比例为1∶1和1∶5时可以产生转移接合子,而当菌体数量比例为5∶1时不能产生转移接合子.共筛选了117株转移接合子,获得了3株不产细菌素的MI15突变株,用光敏生物素标记的Tn5DNA探针与突变株染色体DNA进行斑点杂交,3株突变株均呈阳性反应,证明Tn5转座子已插入到MI15菌株的染色体上引起插入突变并抑制了细菌素的产生.

Pseudomonas aeruginosa PAO1826 containing suicide plasmid pMO75::Tn5 and E.coli S171 containing suicide plasmid pSUP2021::Tn5 were mated with Agrobacterium radiobacter MI15 respectively.The Kanr Rifr transconjugants were obtained at the donor and receptor cell number ratio 1∶1 and 1∶5. But no transconjugants were obtained when the ratio was 5∶1. Three mutants disable to produce bacteriocin were screened from 117 conjugants MI15. Dot blot assays showed that three mutants chromosome DNA were positively hybridized with photobiotin labeled Tn5 DNA probe.It indicated that Tn5 inserted into chromosome of three mutants.