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

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pU8600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pHZ1276。pHZ1276通过接合转移导入变铅青链霉菌(Streptomyces lividans)，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 Application and Construction of a Vector Conjugating and Integrating into Streptomyces and Express Vitrescilla Globin Gene

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.