过表达根瘤血红蛋白基因对活跃链霉菌那西肽产量的影响  

Effects of Overexpressing Hemoprotein Gene from Sinorhizobium meliloti on Nosiheptide Yield of Streptomyces actuosus

作　　者：莫秋燕[1] 宋元达[1] 

机构地区：[1]江南大学食品学院食品与健康研究中心,江苏无锡214122

出　　处：《安徽农业科学》2016年第24期157-159,162,共4页Journal of Anhui Agricultural Sciences

摘　　要：[目的]采用过表达根瘤血红蛋白基因改善活跃链霉菌的溶氧表达,以提高那西肽的产量。[方法]通过全基因组合成的方法得到根瘤菌血红蛋白基因(sm),连接到整合型质粒p IB139上,构建表达载体p IB139-sm。通过大肠杆菌-链霉菌结合转移的方法将sm基因整合到活跃链霉菌基因组上,通过PCR验证各转化子目的片段已整合到活跃链霉菌基因组上,并进行大量筛选得到重组菌株。[结果]经SDS-PAGE和CO结合差示光谱分析,结果显示,目的蛋白均已成功表达且具有相应的蛋白活性。摇瓶培养结果显示,Smf Hb表达对菌体生长和那西肽产量有明显的促进作用,过表达Smf Hb的重组菌株那西肽产量达1 245.7μg/m L,比原始菌株提高39.6%。[结论]过表达sm基因提高那西肽产量的作用优于vhb。[Objective] To improve the dissolved oxygen expression of Streptomyces actuosus by overexpressing hemoprotein Gene from Sinorhi-zobium meliloti, and to enhance the yield of nosiheptide.[ Method] Sinorhizobium meliloti hemoglobin gene （ sm） was obtained through the whole gene synthesis method, which were cloned into the plasmid pIB139 to construct expression vectors pIB139-sm.The plasmid containing sm was introduced into Streptomyces actuosus through conjugal transfer between E.coli and Streptomces.The transformants were verified by PCR, and then were purified by several cycles of single-colony screen.[ Result] The expression and activity of SmfHb was confirmed by SDS-PAGE and CO-difference spectrum analysis, respectively.Flask culture experiment results showed that the nosiheptide yield in SmfHb -overex-pressing strain （1 245.7 μg/mL） was increased by 39.6% （ P〈0.05） compared with the control strain （892.5 μg/mL）.[Conclusion] Overexpressing sm gene is superior to vhb in enhancing the yield of nosiheptide yield.

关 键 词：根瘤菌血红蛋白 活跃链霉菌 那西肽 

分 类 号：S188[农业科学—农业基础科学]