RNA标准物病毒样颗粒表达系统的构建及其特性研究

摘要

为了构建RNA病毒样颗粒（virus-like particles ，VLP）表达系统pET28a（＋）-MS，将插入的外源基因诱导表达获得相应的RNA标准物质病毒样颗粒，将MS2噬菌体包膜蛋白和成熟蛋白基因插入表达质粒pET-28a（＋），构建pET28a（＋）-MS重组子，将带有包装位点序列的外源 RNA对应的 cDNA插入成熟蛋白基因的下游，经过诱导表达含外源 RNA病毒样颗粒。结果表明，pET28a（＋）-MS能表达活性的包膜蛋白（49．8 ku）和成熟蛋白（14．4 ku），能将外源基因 RNA包装成病毒样颗粒，该 VLP长度约1500 bp， RT-PCR反应性优良，能耐受RNase的降解，在血清中37℃经1个月未有明显降解。%To construct virus-like particles(VLP)expression platform pET28a(+)-MS,the inserted foreign gene is induced to express the VLP containing corresponding RNA standard material.The envelope and mature protein genes of MS2 bacteriophages were inserted into plasmid pET-28a(+).The cDNA corre-sponding an exogenous RNA with package sequence was inserted into mature protein gene downstream, and the recombinant pET28a(+)-MS was induced to express exogenous RNA containing VLP.The results showed that,pET28a(+)-MS can express active envelope protein (49.8 ku)and mature protein (14.4 ku),the foreign RNA was packaged into VLP.the VLP is about 1 500 bp long,RT-PCR has excellent re-activity,can withstand the degradation by RNase in serum 3 7 ℃ 1 month and no significant degradation de-tected.