将透明颤菌血红蛋白基因(vgb)转移到链霉菌染色体整合载体的构建  

The Application and Construction of a Vector Conjugating and Int egrating into Streptomyces and Express Vitrescilla Globin Gene

作　　者：张桂敏[1,2] 周秀芬[1] 邓子新[1] 庄永红[3] 

机构地区：[1]华中农业大学生命科学技术学院 [2]湖北大学生命科学学院,武汉430062 [3]湖北大学生命科学学院

出　　处：《华中农业大学学报》2004年第6期602-605,共4页Journal of Huazhong Agricultural University

基　　金：国家自然科学基金项目 (3 0 170 179)资助

摘　　要：链霉菌抗生素的生物合成对培养环境中氧的供应相当敏感 ,为了改善这一溶氧问题 ,本研究通过将透明颤菌血红蛋白基因 (vgb)克隆到含ФC31int,attP基因的诱导型表达载体pIJ86 0 0上 ,构建了可接合转移到链霉菌的整合型表达vgb基因的质粒 pHZ12 76。pHZ12 76通过接合转移导入变铅青链霉菌 (Streptomyceslivi dans) ,所得重组子经Southern杂交验证后 ,进行诱导表达 ,菌体经破碎后所得蛋白粗提物经Western杂交和CO结合试验表明vgb基因在该菌中表达出了有活性的VHb蛋白。The syntheses of antibiotics in Streptomyces are v er y sensitive to the level of dissolved oxygen in fermentation media. Low dissolve d oxygen decreases yield of antibiotics. A Vitrescilla globin gene(vgb) was inserted into pIJ8600, which was a plasmid for the inducible expression vec tor. The recombinant plasmid was named pHZ1276 and introduced into S. lividans through conjugation.After recombinants were confirmed by Southern blotting, w hich demonstrated that this vgb gene has integrated into chromosome of S. lividans , expression of vgb gene were demonstrated by Western blotting an d carbon monoxide binding analysis. Western blotting and carbon monoxide binding analysis of the produced proteins of the recombinant strain showed that active VHb protein has been expressed in S. lividans.

关 键 词：透明颤菌血红蛋白基因 变铅青链霉菌 整合型 基因表达 

分 类 号：Q786[生物学—分子生物学]