摘要
旨在有效检测狂犬病毒抗原及抗体。以狂犬病病毒aG株为模板,反转录扩增糖蛋白膜外区基因,并进行序列测定。将目的片段与表达载体pET-32a(+)分别双酶切、胶回收后相连并转化至Rosseta(DE3)菌株。选取阳性克隆经IPTG诱导表达后,进行SDS-PAGE和Western blotting及质谱鉴定。结果显示,在大肠杆菌中成功表达了糖蛋白膜外区。本研究为建立检测狂犬病毒抗原及抗体的ELISA诊断方法奠定了基础。
The gene of glycoprotein extracellular domain from rabies virus aG strain was amplified by reverse transcriptase-polymerase chain reaction ( RT-PCR ). The product was purified and subcloned into the expression vector pET-32a ( + ). The recombinant plasmid pET-G was transformed into expression strain E. coli Rosetta ( DE3 ) cells and then Isopropyl-J3-D-Thiogalactopyranoside ( IPTG ) was added to induce protein expression. The recombinant protein of glyeoprotein extracellular domain was analyzed by SDS-PAGE, Western blotting and mass spectrometry. Results showed that the recombinant protein had been successfully expressed in E. coll. The study establishes the foundation of ELISA kits to detect the antibodies against rabies virus or antigens of rabies virus.
出处
《生物技术通报》
CAS
CSCD
北大核心
2012年第9期114-119,共6页
Biotechnology Bulletin
基金
兰州市生物医药专项(2011-1-61)
关键词
狂犬病病毒
糖蛋白膜外区
原核表达
Rabies virus Glycoprotein extracellular domain Prokaryotic expression