黑暗链霉菌<i>tobS1</i>-<i>C</i>基因缺失突变株的构建

黑暗链霉菌tobS1-C基因缺失突变株的构建

基金项目: 国家自然科学基金资助项目（No.31070093);国家“重大新药创制”科技重大专项资助项目（No.2012ZX09201-101-008)

摘要: 通过敲除妥布霉素生物合成基因tobS1-C，定向选育阿泊拉霉素高产菌株。PCR扩增tobS1-C基因上下游同源序列和红霉素抗性基因ermE,构建穿梭载体pSH6，利用接合转移法转化黑暗链霉菌Tt49，经同源重组，敲除tobS1-C两基因序列，共2484 bp，获得框内删除突变株黑暗链霉菌T103(△tobS1-C)。发酵验证表明突变株不再合成妥布霉素，只合成阿泊拉霉素。
- 黑暗链霉菌 /
- 阿泊拉霉素 /
- 同源重组 /
- 基因缺失
Abstract: In order to breed a mutant strain for high production of apramycin，carbamoyltobramycin biosynthesis genes tobS1-C were delected from Streptomyces tenebrarius through homologous recombination.Gene deletion vector pSH6 was constructed and introduced into S.tenebrarius Tt49 through conjugal transfer.tobS1-C genes in the chromosome were displaced by deletion allele on the plasmid via double crossover.Then the mutant S.tenebrarius T103（△tobS1-C）was proved to be genetically stable.Shaking flask experiments and TLC analysis showed that the mutant strain only produced apramycin.
- Streptomyces tenebrarius /
- apramycin /
- homologous recombination /
- gene deletion