MOLECULAR CLONING AND CHARACTERIZATION OF RETROVIRUSES ASSOCIATED WITH AIDS

艾滋病逆转录病毒基因组之间的异质性已被清楚地证明。 我们已经使用了分子克隆，限制性内切酶分析， 测序技术来表征人类的分离株 免疫缺陷病毒（HIV）。这些研究可以提供关于 它们的结构和地理多样性， 不同HIV分离株之间的抗原变异和致病性。一个 非洲分离株（扎伊尔-2）和北美分离株（NY-5）的HIV-1， 使用λ噬菌体载体进行分子克隆。每个克隆体都包含 单个LTR（9.2kb基因组），并且在将λ克隆转染入 淋巴细胞，产生感染性病毒颗粒。反向 转录酶（RT）活性类似于构建的感染性 克隆（pNL 4 -3），并且在上清液传代到新鲜淋巴细胞上时， 显示与pNL 4 -3相似的RT动力学。核苷酸全序列 现在已经分析了这些感染性分子克隆中的每一个， 试图制备用于中和试验的每种分离物的抗血清， 进行中。 来自感染HIV-1的T细胞的T细胞克隆（ACH-2）在体外产生HIV-1。 对佛波醇肉豆蔻酸乙酸酯（PMA）刺激的反应。该分离株 被分子克隆，并在转染到淋巴细胞后， 病毒;然而，仅在PMA存在下。新鲜淋巴细胞传代 在不存在诱导PMA的情况下通过RT监测产生的病毒。 目前，我们正在研究PMA在病毒诱导中的生物学作用， HIV-1与病毒感染潜伏期的关系

Heterogeneity among AIDS retroviral genomes has been clearly demonstrated. We have used molecular cloning, restriction enzyme analysis, and nucleotide sequencing techniques to characterize isolates of the human immunodeficiency virus (HIV). These studies may provide data regarding their structural as well as geographic diversity and, possibly, the degree of antigenic variation and pathogenicity among different HIV isolates. An African isolate (Zaire-2) and a North American isolate (NY-5) of HIV-1 were molecularly cloned using lambda bacteriophage vectors. Each clone contained a single LTR (9.2 kb genome) and, upon transfection of lambda clones into lymphocytes, infectious virus particles were produced. Reverse transcriptase (RT) activity resembled that of the constructed infectious clone (pNL4-3) and, upon passage of supernatants onto fresh lymphocytes, displayed similar RT kinetics of pNL4-3. The complete nucleotide sequence of each of these infectious molecular clones has now been analyzed, and attempts to prepare antisera of each isolate for neutralization tests are in progress. A T cell clone (ACH-2) from T cells infected with HIV-1 produced HIV-1 in response to stimulation with phorbol myristic acetate (PMA). This isolate was molecularly cloned and, upon transfection into lymphocytes, produced virus; however, only in the presence of PMA. Passage onto fresh lymphocytes produced virus as monitored by RT in the absence of the inducing PMA. Presently, we are studying the biological role of PMA in viral induction of HIV-1 and the relationship of latency in viral infection.