糖多孢红霉菌多拷贝表达载体pZM的构建

对糖多孢红霉菌染色体上红霉素生物合成基因进行改造 ,已经合成了多种红霉素类似物。在糖多孢红霉菌中对红霉素类似物进行结构修饰 ,以pWOR1 0 9质粒为基础构建糖多孢红霉菌多拷贝表达载体pZM。pZM载体带有PermE启动子、fd终止子、多克隆位点、硫链丝菌肽和氨苄青霉素抗性基因、以及在大肠杆菌和糖多孢红霉菌中复制的ColE1ori和pJV1ori复制子 ,系可在大肠杆菌和糖多孢红霉菌中扩增的穿梭质粒。在糖多孢红霉菌中 ,pZM可以表达氨普霉素抗性基因和绿色荧光蛋白基因 ,从糖多孢红霉菌中提取的表达质粒酶切图谱与转化前一致 ,表明pZM是糖多孢红霉菌中多拷贝、稳定的表达载体。

Construction of Saccharopolyspora erythraea Multi-copy Expression Vector pZM

Based on changing erythromycin biosynthesis genes in Saccharopolyspora erythraea chromosome,many erythromycin analogs had been synthesized.Modifying the structures of erythromycin analogs in Saccharopolyspora erythraea, multi copy expression vector pZM of Saccharopolyspora erythraea had been constructed on base of pWOR109.The vector pZM contained Perm E promoter, fd terminator,multiple clone site,thiostrepton and ampicillin resistance genes,and ColE1 ori and pJV1 ori replicons which made the vector replicate in Escherichia coli and Saccharopolyspora erythraea respectively,and the vector is a shuttle plasmid.In Saccharopolyspora erythraea, pZM could express reporter genes apramycin resistance gene and green fluorescin gene,and the expression plasmid extracted from Saccharopolyspora erythraea had the same enzymatic map with the plasmid before transformation.That shows pZM is a multi copy and stable expression vector in Saccharopolyspora erythraea. ;