摘要
目的:高水平表达新城疫病毒(Newcastle Disease Virus,NDV)标准强毒F48E8株的F基因和HN基因。方法:利用PCR方法扩增出新城疫病毒(Newcastle Disease Virus,NDV)标准强毒F48E8株的去掉N端信号肽和C端跨膜区的F基因和HN基因并将其克隆到原核表达载pET30a上,得到重组质粒rpET30a-NDV/F和rpET30a-NDV/HN,转化到受体菌Rosetta-gamiTM2感受态细胞中,经IPTG诱导表达,SDS-PAGE电泳和Western blot检测。结果:表达的F蛋白大小约为40kD,HN蛋白大小约为51kD左右,符合预期结果。Western blot分析表明,能与抗His-tag单克隆抗体发生特异性反应。结论:重组蛋白在原核表达系统内得到了高水平的表达。说明了去除F和HN基因N端信号肽和C端跨膜区的必要性。
Objective： To high effective express fusion （F） and haemaglutinin neuraminidase germ （HN） of Newcastle Disease Virus （NDV） stan- dard velogenic F48E8 strain. Method： The F germ and HN gene which N-terminal signal peptide and C - terminal transmembrane domain were deleted were amplified by PCR and cloned into the multiple cloning sites of pET30a vectors. The recombinant plasmids rpET30a - NDV/F and rpET30a- NDV/HN were identified and transformed into Rosetta- gami^TM2 competent cells and induced by IPTG. The recombinant proteins were proven by SDS- PAGE and Western blot assay. Result： The F protein and HN protein of NDV were 40kD and 51kD in size respectively, according with anticipates. The results of Western blot showed that these proteins were specifically reacted with His - tag moneclonal antibody. Conclusion： These results indicate that the recombinant protein had high effective express and getting rid of N - terminal signal peptide and C - terminal transmembrane domain were necessary.
出处
《生物技术》
CAS
CSCD
2008年第3期11-13,共3页
Biotechnology
