Ketogulonigenium vulgare四环素诱导表达穿梭质粒的构建  

Construction of an Inducible Expression Shuttle Vector for Ketogulonigenium vulgare Allowing Tightly Regulable Gene Expression

作　　者：罗婉月 李天明[1] 于莹[1] 许湄雪 仪宏[1] 

机构地区：[1]河北科技大学,石家庄050018

出　　处：《中国生物工程杂志》2015年第5期81-86,共6页China Biotechnology

摘　　要：采用重叠延伸PCR方法合成阻遏蛋白的编码基因tetr和插入操纵基因teto的Ketogulonigenium vulgare山梨糖脱氢酶启动子psndhteto的基因序列,借助宽宿主质粒p BBR1MCS-5,构建四环素诱导表达的穿梭质粒,转化Ketogulonigenium vulgare,获得阳性重组菌株,实现卡那霉素抗性的调控表达,结果表明:培养重组菌株2小时后,添加0.4μg/ml的四环素诱导剂后,能够在含有卡那霉素的培养基中生长,不添加四环素诱导剂的重组菌株不能在含卡那霉素的培养基中生长,确定了最适四环素的诱导浓度为0.6μg/ml。The genes encoding tet repressor proteins and Ketogulonigenium vulgare sorbose dehydrogenase operator which inserted operon teto were synthesized by overlapping PCR. The shuttle vector of tetracycline induced expression with broad host plasmid pBBR1MCS-5 was constructed and transformed Ketogulonigenium vulgare. The resistance of kanamycin was regulatively expressed after getting the positive recombinant strains. The results show that the cultivation of recombinant strains could grow in the medium containing kanamycin where added 0.4mg/ml tetracycline inducers after 2 hours, while they can＇ t grow in the medium containing kanamycin where didn＇ t add tetracycline inducers. The optimal induction of tetracycline concentration determined was 0.6mg/ml.

关 键 词：生酮古龙酸杆菌 基因合成 穿梭质粒 四环素诱导 

分 类 号：Q789[生物学—分子生物学]