嵌合人牛免疫缺陷病毒cDNA的构建及其在MT_4细胞中的活性分析

Construction and analysis of activity of an HIV-1/bovine immunodeficiency virus chimeric clone cDNA

目的 构建嵌合人 牛免疫缺陷病毒 (HBIV)cDNA ,研究牛免疫缺陷病毒BIVTat及LTR在人源MT4 细胞中的活性。方法 PCR扩增BIV的tat、LTR及HIV的gag、pol、env片段 ,定向插入到pBluescriptSK(+)载体上 ,嵌合质粒转染MT4 细胞 ,逆转录聚合酶链反应 (RT PCR)检测嵌合病毒中的基因转录 ,反转录酶活性测定嵌合病毒基因的表达。结果 BIVtat、HIVgag在MT4 细胞中均已得到转录 ,并已合成具有生物学活性的反转录酶。结论 在人源MT4 细胞中 ,嵌合病毒的BIVLTR具有启动子活性 ,BIVTat具有反式激活功能。

Objective Chimeric human/bovine immunodeficiency virus (HBIV) cDNA was constructed by replacing HIV tat and LTR with bovine immunodeficiency virus (BIV) tat and LTR to study the activity of BIV tat and LTR in the chimerae Methods The target fragments of BIV tat, LTR and HIV gag, pol, env were respectively amplified by using PCR and sequentially inserted into pBluescript SK(+) vector The chimeric clone was transfected into human MT 4 cells The transcript and gene expression of the HBIV chimeric virus were detected by using RT PCR and a reverse transcriptase assay, respectively Results BIV tat mRNA and HIV gag mRNA were detected The reverse transcriptase activity of the chimeric virus was analyzed in the fluctuation curve Conclusion In chimeric HBIV cDNA transfected MT 4 cells, BIV tat and HIV gag were transcripted The reverse transcriptase of the chimeric virus had biological activity These data suggest that in MT 4 cells, BIV LTR had promoter activity and BIV tat had the function of transactivation in the chimeric virus The study of the chimeric virus with infectivity is in progress