【摘要】 用共转化技术分别将两种亚型的单拷贝HBV-DNA重组质粒与HSV-tk基因一起导入Ltk细胞获得成功。6系克隆化的tk~+细胞分泌HBsAg。分泌量较高的pA-3及PC-3两系细胞分泌物为特异性HBsAg,并含有约22nm大小的颗粒性HBsAg。初次细胞克隆HBsAg表达不稳定,再次克隆获得PC-3-1亚系传代160天仍表达HBsAg,说明再次克隆是选择HBsAg表达较稳定的细胞系的必要手段。单拷贝HBV-DNA转化成功,似说明首尾相连的双拷贝HBV-DNA并非必需。更多还原

【Abstract】 Two plasmids containing a single copy of hepatitis B virus (HBV) genome (ayw or adw) have been introduced into mouse Ltk-cells by calcium precepitation and cotransformation with cloned herpes simplex virus thymidine kinase (tk) gene. Under the selective pressure of HAT medium, 6 cloned tk~+ cell lines were found to excrete specific HBsAg in cell culture medium. Two cell lines, pA-3 and pC-3 were selected for further studies. These cell lines were found to be composed of HBsAg positive and HBsAg negative cells on subcloning. The pC-3-1 subline cells obtained from pC-3 cell line by subcloning retained their ability to excrete HBsAg after 16o days passage. This phenomenon shows. that the subcloning procedure is essential for obtaining stable cell lines continously excreting HBsAg. The HBsAg excreted into the cell culture medium was shown by immunoelectromicroscopy to be composed of particles about 22nm diameters, being similar to those found in hu- man serum. Since in our experiments, cotransformation was successful with a single copied HBV-DNA, it seems to indicate that two copies of HBV-DNA in tandem are not essential.更多还原