用美国白蛾核型多角体病毒表达家蚕促前胸腺激素基因

携带有家蚕促前胸腺激素PTTH基因的重组转移载体pHcMU1PTTH(含有麻蝇毒素IA信号序列 )与美国白蛾核型多角体病毒 (HcNPV)基因组DNA共转染SPIM细胞 ,经 2轮空斑纯化 ,筛选出重组病毒HcNPVPTTH+ ,SDSPAGE和Westernblot分析表明 :在细胞培养上清中可检测到PTTH的存在 ,说明麻蝇毒素IA信号序列可被SPIM细胞识别。用抗PTTH的兔血清进行Westernblot分析可显示分子量为2 0ku左右的 2条带 ,但当培养液内加入衣霉素时 ,仅出现 1条分子量小于 1 8ku的信号带 ,推测 2条信号带的产生与糖基化修饰不一致有关

Bombyx mori Prothoracicotropic Hormone Gene Was Expressed by Recombinant Hyphantria cunea Nuclear Polyhedrosis Virus

After two times plaque assay, the recombinant virus HcNPV PTTH was screened by cotransfecting the recombinant transferring vector pHcMU 1 PTTH contained Bombyx mori prothoracicotropic hormone (PTTH) gene (with sarcotoxin IA signal sequence) and Hypantria cunea nuclear polyhedrosis virus (HcNPV) genomic DNA into SPIM cell.SDS PAGE and Western blot analysis indicated that the PTTH could be detected in SPIM cell culture supernatant and sarcotoxin IA signal sequence recognized in SPIM cell.Two spicific signal bands whose molecular weight were around 20 ku were detected,but only one band whose molecular weight was smaller than 18 ku was revealed when tunicamycin was added into the cell culture media using Western blot analysis with anti rabbit PTTH serum.It is deduced that the difference of glucosylation was involved.