Construction and application of an In-fusion-based high-throughput system to detect HIV phenotypic resistance
WU Shou-li1, 2, LIU Feng2, YAN Ping-ping1, 2, WANG Zheng-hua1, XIE Mei-rong1, YAN Yan-sheng1, 2
1. Fujian Provincial Center for Disease Control and Prevention/Fujian Provincial Key Laboratory of Zoonosis Research, Fuzhou 350001, China; 2. School of Public Health, Fujian Medical University, Fuzhou 350004, China
Abstract：The aim of this study is to develop an In-fusion-based high-throughput HIV novel phenotypic resistance detection system and evaluate its application in the clinical detection of HIV-1 drug resistance. The new HIV back-bone vector pLWJ-SV40-Luc-LacZ was constructed by inserting amplified LacZ gene into pLWJ-SV40-Luc. Using the In-fusion cloning technology, resistance test vectors (RTVs) were constructed by incorporating amplified target genes into pLWJ-SV40-Luc-LacZ and replacing the LacZ fragment. The pseudovirus stocks used for drug susceptibility test were produced by co-transfecting 293T cells with RTVs and a plasmid that provided vesicular stomatitis virus glycoprotein (VSV-G). Viral replication was monitored by measuring luciferase activity in infected target cells. A total of 36 clinical plasma samples from HIV-1-infected humans were tested, and target fragments were successfully amplified from 30 samples (83.3 %) and 26 RTVs were successfully constructed by In-fusion cloning, of which 21 RTV had active infection. The phenotypic profiles of the pseudoviruses from 21clinical samples completely matched the observed genotypes. The results demonstrate that an In-fusion-based high-throughput HIV novel phenotypic resistance detection system was successfully developed, and this testing system appears to be applicable for testing phenotypic resistance of clinical HIV-1 strains and provides information for clinical therapy.
吴守丽, 刘峰, 颜苹苹, 王征桦, 谢美榕, 严延生. 基于In-fusion技术的高通量HIV表型耐药检测系统的建立及应用研究[J]. 中国人兽共患病学报, 2019, 35(4): 285-291.
WU Shou-li, LIU Feng, YAN Ping-ping, WANG Zheng-hua, XIE Mei-rong, YAN Yan-sheng. Construction and application of an In-fusion-based high-throughput system to detect HIV phenotypic resistance. Chinese Journal of Zoonoses, 2019, 35(4): 285-291.
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2019, Vol. 35 
