摘要
pMM2066质粒以EcoR Ⅰ酶切回收ATG前只有12bp的乙肝核心抗原基因片段,将此基因片段插入痘苗转移载体pGJP-5的P7.5启动子后,组建成P5.C2066质粒,通过细胞内同源重组技术,于BUdR存在条件下用人TK-143细胞筛选出三株能表达HBcAg的重组痘苗病毒株。感染细胞上清液1:64稀释时仍能用ELISA法测到HBcAg活性,同时也能测到滴度相近的HBeAg活性。免疫电镜可见平均为26.6nm的HBcAg颗粒。进行了不同量的重组痘苗病毒感染不同细胞,和不同培养温度对HBcAg表达影响的观察。
Plasmid pMM2066 was digested with EcoR I restriction enzyme and the 870 bp fragment which containing 12 bp upstream to the ATG codon and structure gene of C antigen was separated. This DNA fragment was then inserted into the transfer vector pGJP-5 just downstream to the P7.5 promoter. Thus, the recombinant vector P5-C2066 was constructed. Using homologous recombination technique 3 TK- recombinant vaccinia viruses expressing HBcAg were isolated by plaque assay in TK-143 cell in the presence of BUdR. Up to 1:64 dilution of supernatant of cells infected with recombinant virus still showed significant positive results by using ELISA method. About the same titer of the HBeAg activity was also detected. 26.6 nmHBcAg particles were visualized by electron microscopy. In addition, the effects of M.O.I, kinds of cell cultures and euhivated temperature on the expression level of HBcAg had also been investigated.
出处
《生物工程学报》
CAS
CSCD
北大核心
1989年第3期201-206,共6页
Chinese Journal of Biotechnology
基金
中国自然科学基金
