Modification of HIV-1 (Human immunodeficiency virus type one) Chinese epidemic strain CRF01_AE env gene

A gene and modified technology, applied in plant gene improvement, gene therapy, genetic engineering, etc., can solve urgent problems such as AIDS vaccine research

Inactive Publication Date: 2014-02-19
中国疾病预防控制中心病毒病预防控制所 +2
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Problems solved by technology

These data show that although the rate of new HIV infections has been declining, the total number of HIV infections is still increasing, and the situation of the AIDS epidemic is still severe. Research on effective preventive and therapeutic AIDS vaccines is urgent

Method used

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  • Modification of HIV-1 (Human immunodeficiency virus type one) Chinese epidemic strain CRF01_AE env gene
  • Modification of HIV-1 (Human immunodeficiency virus type one) Chinese epidemic strain CRF01_AE env gene
  • Modification of HIV-1 (Human immunodeficiency virus type one) Chinese epidemic strain CRF01_AE env gene

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Embodiment Construction

[0018] 1. Cloning of HIV-1 CRF01_AE env Gene and Acquisition of Consensus Sequence

[0019] For the peripheral venous blood of 100 HIV-1 infected patients in Beijing collected in EDTA anticoagulant blood collection tubes, QIAamp DNA Blood Mini Kit (Cat. No. 51106) from QIAGEN was used to extract proviral DNA according to the instructions. Store at -20°C. The gag gene fragment was amplified by the nested-PCR method, and used for sample HIV-1 type analysis. Using Ex Taq (Cat.No.DRR100A) produced by TaKaRa Company, Gag F2 (5'ATG GGT GCG AGA GCG TCA RTA TTA A 3', HXB2 position 790-814), Gag e2 (5'TCC AAC AGC CCT TTT TCC TAG G 3', HXB2 position 2032~2011) as the outer primer for the first round of PCR reaction, the reaction system is 25μl system: 10×Ex Taq Buffer 2.5μl, 25mM dNTP Mixture 2μl, TaKaRa Ex Taq (5U / μl) 0.125 μl, template 2μl, primers GagF2 and Gag e2 each 0.5μl, add ddH 2 0 to 25 μl. Reaction program: 94°C for 5 minutes, 52°C for 1 minute, 72°C for 2 minutes and 30...

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Abstract

The invention provides a modified CRF01_AE env gene. According to the modified CRF01_AE env gene, the GC content is increased, and an internal inhibitory sequence (INS) with higher AU content is damaged, so that the gene is more suitable for being expressed in a eukaryotic cell. The gene can be expressed efficiently in the eukaryotic cell without depending on regulatory protein Rev and a response element of the regulatory protein Rev; compared with the gene before modification, the expression quantity is remarkably increased; the modified mod.AE env gene is selected as a vaccine gene; and if HIV-1 vaccines are restructured, the immunogenicity of the vaccines can be improved possibly.

Description

technical field [0001] The invention belongs to the field of biomedicine. The present invention relates to the acquisition and transformation of the env region consensus sequence of the Chinese representative HIV-1 CRF01_AE subtype strain to improve its expression level. Background technique [0002] Human immunodeficiency virus type one (HIV-1) is the main pathogen of acquired immunodeficiency syndrome (AIDS). The latest UNAIDS report estimates that there are about 34 million HIV-infected people worldwide, of which women and children account for about 55%, as many as 18.4 million, and nearly 2 million infected people die of ADIS-related diseases every year. Although extensive publicity and education, behavioral interventions, and treatment interventions have reduced the HIV infection rate and mortality to a certain extent in the past decade, the number of new infections is still high at more than 2 million per year. By the end of 2011, there were about 780,000 HIV-infecte...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/49C07K14/16C12N15/85C12N5/10A61K48/00A61K39/21A61P31/18C12R1/93
CPCY02A50/30
Inventor 曾毅冯霞陈丹瑛何小周余双庆李泽琳
Owner 中国疾病预防控制中心病毒病预防控制所
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