摘要
ＩＪ６０２１和ｐＩＪ４１２３是链霉菌的高拷贝表达载体，它们携带有受硫链丝菌素诱导的强启动子ＰｔｉｐＡ。分别在它们的合适位点插入大肠杆菌质粒的复制子和在大肠杆菌中选择用的抗性标记基因（ｂｌａ），得到了两个能在大肠杆菌和链霉菌中穿梭复制、并保持结构稳定的链霉菌表达载体：ｐＨＺ１２７１和ｐＨＺ１２７２。将透明颤菌（Ｖｉｔｒｅｏｓｃｉｌｉａｓｐ．）血红蛋白基因（ｖｈｂ）克隆到ｐＨＺ１２７２中，用它转化变铅青链霉菌（Ｓｔｒｅｐｔｏｍｙｃｅｓｌｉｖｉｄａｎｓ），经Ｗｅｓｔｅｒｎｂｌｏｔｉｎｇ分析和ＣＯ结合实验表明，在变铅青链霉菌中表达出了有生物活性的透明颤菌血红蛋白，从而证明所构建的ｐＨＺ１２７２载体具有在链霉菌中表达外源基因的功能。
pIJ6021 and pIJ4123 are high copy number vectors for gene expression in Streptomyces .They contain a strong,thiostrepton inducible promoter,P tipA· Two E.coli Streptomyces shuttle vectors,pHZ1271 and pHZ1272,were constructed by inserting a replicon and bla gene from E.coli into pIJ4123 and pIJ6021 respectively.Both vectors were structurally stable either in E.coli or in Streptomyces lividans. To demonstrate the utility of pHZ1272,the hemoglobin gene of Vitreoscilla (vhb) was cloned into pHZ1272 and expressed in Streptomyces lividans. The expression product (Vhb) was detected by Western blotting and carbon monoxide binding activity analysis.
出处
《生物工程学报》
CAS
CSCD
北大核心
1998年第1期6-12,共7页
Chinese Journal of Biotechnology
基金
国家自然科学基金
国家教委资助
关键词
链霉菌
大肠杆菌
穿梭载体
血红蛋白
基因表达
Streptomyces lividans, E.coli ,shuttle vectors, vhb gene expression
