摘要
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一个通用载体平台。方法将MS2噬菌体基因组中包括的成熟酶蛋白和包膜蛋白的基因编码序列的1.7kbcDNA目的片段,用HindⅢ和EcoRI酶切后,与用相同内切酶酶切的表达载体质粒pET28b,在T4DNA连接酶的存在下连接,构建一新的表达载体pINCCL,再转化BL21-DE3E.Coli进行原核表达。结果成功构建得到了新的表达载体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,可作为一个耐RNase的RNA标准品和质控品的构建和制备表达通用载体平台,将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 MS2 phage RNA genome, which encodes coat protein and maturase protein, and expression vector pET28b DNA are ligated together with T4 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 successf ully. 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 a express plasmid carrier platform that can be in common use for preparation of RNase -resistant RNA standards and controls.
出处
《医学检验与临床》
2008年第5期33-35,共3页
Medical Laboratory Science and Clinics
