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目的观察急性感染期艾滋病病毒I型(HIV-1)gp160的全长基因序列及感染特征。方法从一例处于Feibig I期HIV-1感染者血浆中提取RNA,扩增gp160全长基因并测序,分析其生物学信息;将gp160全长基因与pcDNA3.1His/V5真核表达载体连接,构建Env-pcDNA3.1真核表达质粒,与骨架质粒pNL4-3.Luc.R-E-共转染293细胞,包装出假病毒。用包装的假病毒感染ghost细胞,测定感染细胞的荧光素酶活性(RLU),鉴定假病毒的感染活性。结果成功扩增出gp160全长基因,嗜性预测为CCR5,N-糖基化位点数与标准株HXB2相同,但gp120糖基化程度更高,氨基酸变异主要集中在V1~V5区。假病毒感染试验显示,RLU值达到7log。结论获得了处于急性感染期的HIV-1gp160基因序列和高感染活性的假病毒。
Objective To observe the full length gene sequence of HIV-1 gp160 and its infection characteristics during acute infection. Methods RNA was extracted from the plasma of an HIV-1 infected Feibig patient I, and the full-length gp160 gene was amplified and sequenced to analyze its biological information. The full-length gp160 gene was linked to the eukaryotic expression vector pcDNA3.1His / V5 to construct Env-pcDNA3.1 eukaryotic expression plasmid was co-transfected with the backbone plasmid pNL4-3.Luc.RE-293 into 293 cells to package pseudovirions. The ghost cells were infected with the packaged pseudovirus, the luciferase activity (RLU) of the infected cells was determined, and the infection activity of the pseudovirus was identified. Results The full-length gp160 gene was successfully amplified and its tropism was predicted to be CCR5. The number of N-glycosylation sites was the same as that of the standard strain HXB2, but the degree of glycosylation of gp120 was higher. The amino acid variations mainly concentrated in V1 ~ V5. Pseudovirus tests showed that the RLU reached 7 logs. Conclusion The HIV-1 gp160 gene sequence and the highly infectious pseudovirus in the acute phase were obtained.