bldD基因过量表达对红色糖多孢菌红霉素产量及孢子形成的影响

目的通过增加红色糖多孢菌调控基因bldD拷贝,获得红霉素高产菌株,并探讨bldD基因过量表达对红色糖多孢菌形态分化的影响。方法以红色糖多孢菌A226为出发菌株,通过染色体同源重组方法获得红色糖多孢菌A226bldD基因失活突变菌株A226-△bldD。将bldD基因表达质粒pZMW-bldD分别导入突变株A226-△bldD及出发菌株A226中,获得A226-△bldD/bldD菌株和A226/bldD菌株,利用TLC法和HPLC法对其发酵产物进行分析,并观察孢子生长状况。结果红色糖多孢菌A226中bldD基因失活后产红霉素水平明显降低,不能形成孢子;bldD基因的回复表达使A226-△bldD突变菌株产红霉素能力得到恢复,且孢子形成恢复正常;在红色糖多孢菌A226中过量表达bldD基因可提高红霉素产量,较出发菌株A226提高20%,同时可使孢子形成时间提前。结论在红色糖多孢菌中过量表达bldD基因,可获得红霉素高产菌株,同时会影响红色糖多孢菌形态分化进程。

Effect of bldD gene overexpression on erythromycin output and spore formation of Saccharopolyspora erythraea

Objective To obtain an erythromycin high-yield strain by increasing expression of bldD gene in Saccharopolyspora erythraea,and explore the effect of overexpressing bldD gene on the morphological differentiation of Sac.erythraea.Methods Based on Sac.erythraea A226 strain,a mutant A226-△bldD with bldD gene disrupted was constructed by chromosomic homologous recombination method.When expression plasmid pZMW-bldD was introduced into Sac.erythraea A226-△bldD and A226,respectively,A226-△bldD/bldD and A226/bldD strains were obtained.Bacterial suppression test,TCL and HPLC were used to analyze the fermentation products of A226-△bldD/bldD and A226/bldD,and the spore formation process of both strains was observed.Results When the bldD gene was disrupted in Sac.erythraea A226,the obtained Sac.erythraea A226-△bldD produced a lower erythromycin yield,and no spores were formed,while introducing bldD gene into A226-△bldD restored the ability to produce erythromycin and to form spores.It was showed that when bldD gene was introduced into A226,the erythromycin yield increased by 20% and the spore formation occurred earlier compared with Sac.erythraea A226.Conclusion High-yield erythromycin strain of Sac.erythraea could be obtained by overexpressing bldD gene in Sac.erythraea,and adding a bldD gene copy could advance the time of morphological differentiation in Sac.erythraea.