大肠杆菌中人心钠素基因在P_L和T7启动子控制下的高效表达

Expression of Atrial Natriuretic Peptide Gene under the Control of P_L and T7 Promoter in E. coli

将Ｐ１噬菌体的ｒｅｆ基因与人心钠素的嵌合基因ＲＥＦ－ＡＮＰ分别克隆入带Ｐ＿Ｌ启动子和Ｔ７启动子的两种表达载体，获得高效表达重组质粒ｐＺＪＭ１和ｐＺＪＭ４．ｐＺＪＭ１转化ＤＨ５α、ｐＺＪＭ４转化ＨＭＳ１７４，在表达细菌培养到ＯＤ＿（６００）＝０．２５时分别进行温度诱导和化学诱导，快速凝胶扫描结果表明ＲＥＰ－ＡＮＰ表达量各占细菌可溶性总蛋白的６７．１％和２８．１％；在ＯＤ＿（６００）＝１．０或２．０时进行诱导，均能维持高效表达；且温度诱导的表达效率总是优于化学诱导。

The chimeric gene was cloned into two expression vectors and two re-combinant plasmids were obtained.One recombinant plasmid called pZJM1,containing P_Lpromoter,can be expressed by temperature induction. The other one,pZJM4 which containsT7 promoter,can be expressed by chemical induction. pZJM1 and pZJM4 were transformedinto E. coli strains DH5α and HMS174 respectively.Induction of expression started when thebacteria grew at OD_(600)=0. 25 and REF-ANP fusion protein wsa obtained with a yield cor-responding to 67.1%for pZJM1/DH5α and 28.1%for pZJM4/HMS174 of total cellular solu-ble proteins estimated by rapid gel scanning on SDS/PAGE,In addition,expression of thefusion protein,either by temperature induction or by chemical induction,can maintain highlevel when the induction started at high OD_(600) values(OD_(600)=1. 0 or 2.0)of bacterial cul-ture.