耐核糖核酸酶病毒样颗粒的构建和表达

Construction and expression of RNase-resisting virus-like particles

目的 为耐核糖核酸酶 (RNase)的RNA标准品和质控品的表达制备提供 1个通用载体平台。方法 将MS2 噬菌体基因组中成熟酶蛋白和包膜蛋白基因编码序列的 1 7kbcDNA目的片段 ,用HindⅢ和EcoRⅠ酶切后 ,用于相同内切酶酶切的表达载体质粒pET2 8b中 ,并在T4 DNA连接酶的存在下连接 ,构建一新的表达载体pINCCL ,再转化BL2 1 DE3E Coli进行原核表达。结果 成功构建出新的表达载体pINCCL ,经原核表达为耐RNase的病毒样颗粒。结论 本研究得到的pINCCL表达载体及原核表达系统 ,可作为 1个耐RNase的RNA标准品与质控品的构建和制备表达通用载体平台 ,以促进有关标准品和质控品的研究。

Objective To provide a express plasmid carrier platform that can be in common use for preparation of RNase-resistant RNA standards and controls. Methods A cDNA fragment of MS 2 phage RNA genome, which encodes coat protein and maturase protein, and expression vector pET28b DNA are ligated together with T 4 DNA ligase after digested with HindⅢ and EcoR Ⅰ restriction nucleases. Then, a new expression plasmid carrier pI NCCL is contructed。 The prokaryotic expression was carried out by transform pI NCCL into BL21-DE3 E。Coli. Results A new expression plasmid carrier pI NCCL is contructed successfully. RNase-resistant virus-like particles were obtained after prokaryotic expression of pI NCCL. Conclusion The expression plasmid carrier pI NCCL contructed and prokaryotic expression system can be used as a express plasmid carrier platform that can be in common use for preparation of RNase-resistant RNA standards and controls.