反转录病毒介导的稳定表达HIV-1 Gag蛋白和增强型绿色荧光蛋白细胞系的建立

畜牧兽医学报 ›› 2011, Vol. 42 ›› Issue (2): 203-209.doi:

反转录病毒介导的稳定表达HIV-1 Gag蛋白和增强型绿色荧光蛋白细胞系的建立

王婧¹，李昌²，李林溪¹，胡博¹，丛艳昭³，任大勇¹，王卓越¹，杜寿文¹，金宁一¹，²*

1.吉林大学畜牧兽医学院，长春 130062；2.军事医学科学院全军基因工程重点实验室，长春 130062； 3.延边大学农学院，龙井 133400

收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2011-02-25 发布日期:2011-02-25
通讯作者: 金宁一

Continuously Expressing HIV-1 Gag and EGFP by Retroviral Vector InducedSP2/0 Cell Lines

WANG Jing¹, LI Chang², LI Lin-xi¹, HU Bo¹, CONG Yan-zhao³,
REN Da-yong¹, WANG Zhuo-yue¹, DU Shou-wen¹, JIN Ning-yi^1,2*

1.College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, China; 2.Genetic Engineering Laboratory of PLA, Academy of Military Medical Sciences of PLA, Changchun 130062, China; 3. College of Agriculture,Yanbian University, Longjing 133400, China

Received:1900-01-01 Revised:1900-01-01 Online:2011-02-25 Published:2011-02-25

摘要/Abstract

摘要： 本研究旨在建立中国流行株人免疫缺陷病毒（HIV-1）衣壳蛋白(Gag)哺乳动物稳定表达细胞系。将HIV-1 核心蛋白基因gag 和增强型绿色荧光蛋白基因EGFP依次串联插入反转录病毒载体pFB-neo，构建重组反转录病毒载体pFB-gag-EGFP，并与含有辅助病毒gag-pol和env基因的质粒pVPack-GP、pVPack-10A1共转染HEK293T细胞，包装出的反转录病毒感染小鼠骨髓瘤细胞SP2/0。荧光显微镜下观察绿色荧光蛋白EGFP表达，验证HIV-1核心蛋白Gag表达，G418抗性筛选阳性细胞。结果表明，HIV-1核心蛋白Gag和增强型绿色荧光蛋白可在SP2/0细胞中稳定表达，HIV-1核心蛋白gag基因稳定表达细胞系成功建立，为抗AIDS治疗用基因工程制剂及靶向药物的活性检测提供了理想方法。

Abstract: To construct a retroviral vector mediated mammalian cell expression cell line of the human immunodeficiency virus (HIV-1) core protein, the recombinant retroviral vector pFB-gag-EGFP was constructed by inserting core protein gene gag of HIV-1 and enhanced green fluorescent protein gene EGFP into pFB-neo by turns. The recombinant retroviral vector, the pVPack-GP plasmid containing the helper virus gag-pol gene and pVPack-10A1 plasmid containing env gene were cotransfected into packaging cell line HEK-293T with liposome-mediated transduction, and then the recombinant retrovirus was collected and infected SP2/0 cells. The green fluorescent protein was observed under fluorescence microscope, and the expression of core protein Gag of HIV-1 was verified. The resistance cells were screened by G418. The core protein Gag of HIV-1 and green fluorescent protein could express in SP2/0 cells, and the mammalian cell line of the core protein gene gag of HIV1 had been constructed successfully, which laid a mouse cell model of activity assay of anti.AIDS genetic engineering therapeutic agents and targeted drugs.

王婧;李昌;李林溪;胡博;丛艳昭;任大勇;王卓越;杜寿文;金宁一;. 反转录病毒介导的稳定表达HIV-1 Gag蛋白和增强型绿色荧光蛋白细胞系的建立[J]. 畜牧兽医学报, 2011, 42(2): 203-209.

WANG Jing;LI Chang;LI Lin-xi;HU Bo;CONG Yan-zhao;REN Da-yong;WANG Zhuo-yue;DU Shou-wen;JIN Ning-yi;. Continuously Expressing HIV-1 Gag and EGFP by Retroviral Vector InducedSP2/0 Cell Lines[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2011, 42(2): 203-209.

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