重组逆转录病毒载体携带乙型肝炎病毒载体的构建与表达

目的 探索利用逆转录病毒载体表达HBV载体的可能性及复制缺损性HBV能否被正确包装。方法 在逆转录病毒载体中插入表达显性阴性突变体的HBV基因复制单元，制备重组逆转录病毒后分别用以感染Hep G2和2．2．15细胞，免疫荧光法检测细胞内HBV核心抗原的表达，PCR检测上清液中重组HBV颗粒。结果 在包装细胞系PA317中能形成较高滴度的重组逆转录病毒，用以感染Hep G2细胞后，可检出核心抗原的表达。感染2．2．15细胞后，在培养上清液中能检测出突变型HBV颗粒。结论 重组逆转录病毒能携带HBV载体在细胞内表达，并在有野生型HBV提供结构蛋白的前提下，发挥HBV载体的功能，可望用于抗HBV基因治疗。

Construction and expression of recombinant retrovirus vector carrying HBV vector

Objective To explore the possibility of using retrovirus vector to carry HBV vector,and to prove that replication defective HBV could be normally packaged. Methods Two kinds of full length of mutant HBV gene,which express dominant negative mutants,were inserted into retrovirus vector. After recombinant retroviruses were harvested,they were used to infect Hep G 2 and 2.2.15 cell line. Then the expression of HBV core antigen in the Hep G 2 cell was examined by immune fluorescence,and the existence of recombinant HB virion in the culture medium was examined by PCR.Results High titer of recombinant retroviruses were obtained in the culture medium of transfected PA317 cell line.Core antigen was detectable in the recombinant retrovirus infected Hep G 2 cell. Recombinant HB virion was detectable in the culture medium of recombinant retrovirus infected 2.2.15 cell.Conclusion The results suggested that recombinant retrovirus could carry HBV vector and express HBV products.When structural protein is offered by wt HBV,the recombinant retrovirus may function as HBV vector,therefore it could be used in anti HBV gene therapy.