人呼吸道合胞病毒M2-1基因原核表达质粒的构建与表达

Construction and expression of human respiratory syncytical virus M2-1 gene plasmid in prokaryotic cells

目的 构建人呼吸道合胞病毒 (hRSV)M2 1基因原核表达质粒载体 ,使其于原核细胞中表达。方法 利用Hep 2细胞培养hRSV ,提取感染细胞总RNA ,通过RT PCR扩增M2 1基因片段。目的片段与质粒pGEX 2T经EcoRI、XhoI双酶切后 ,连接、转化 ,筛选出阳性克隆。阳性克隆菌经诱导剂诱导后 ,表达M2 1融合蛋白。结果 成功地构建了M2 1基因原核表达质粒载体 ,实现了其于原核中的表达。结论 建立了hRSV基因M2 1原核表达载体 ,为进一步研究hRSV奠定基础。

Objective To construct human respiratory syncytical virus (hRSV) M2 1 gene vector and then express the vector in prokaryotic cells. Methods Total RNA of hRSV infected Hep 2 cells was extracted and M2 1 gene fragment was amplified with reverse transcriptase PCR (RT PCR). Digested by EcoR Ⅰ and Xho Ⅰ, the amplified M2 1 fragment and pGEX 2T were ligated and the product was tranfected to prokaryotic cells. The positive cloned bacteria were indentified with PCR and enzyme digestion and then induced by IPTG to express M2 1 gene. Results The prokaryotic expressing plasmid containing hRSV M2 1 gene was constructed and the target gene was expressed in objective cells. Conclusion The construction of the vector coding for hRSV M2 1 gene will provide a foundation for further study on hRSV.