HIV-1gp120 gene consensus sequence optimized by codon and gp120 nucleic acid vaccine

A HIV-1, codon optimization technology, applied in gene therapy, genetic engineering, plant gene improvement, etc., to achieve a good immune response

Inactive Publication Date: 2010-11-17
王世霞 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The high mutation of Gp120 and the resulting neutralizing ant...

Method used

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  • HIV-1gp120 gene consensus sequence optimized by codon and gp120 nucleic acid vaccine
  • HIV-1gp120 gene consensus sequence optimized by codon and gp120 nucleic acid vaccine
  • HIV-1gp120 gene consensus sequence optimized by codon and gp120 nucleic acid vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1. Analysis of gp120 amino acid sequence of HIV-1AE, BC and ThB subtypes in China

[0039] The gp120 amino acid sequences of known Chinese HIV-1AE, BC and ThB subtypes in the gene bank were analyzed. According to the results of the sequence arrangement and the analysis results of the genetic evolution tree ( figure 1 ), designed the gp120 amino acid consensus sequences of Chinese HIV-1AE, BC and ThB subtypes.

Embodiment 2

[0040] Example 2. Design and synthesis of codons encoding consensus amino acid sequences

[0041] First, use the gene software MacVector 7.2 to analyze the gene sequence encoding the consensus amino acid sequence of the envelope protein gp120 of HIV-1 epidemic strains in China (A / E recombinant subtype, B / C recombinant subtype and ThB subtype), and find out its codons Bias and codon sites in the original sequence where codon usage bias differs from mammalian codon usage bias and from E. coli codon usage bias. For amino acids with the same codon usage preference in mammals and Escherichia coli, the codons encoding the amino acid in the original sequence were replaced by codons favored by both mammals and Escherichia coli, and codon-optimized coding AE, BC and ThB were designed respectively The gene sequences of subtype gp120 proteins are respectively SEQ ID NO.4, SEQ ID NO.5 and SEQ ID NO.6. After codon optimization, the DNA sequence of gp120 gene changed, but the amino acid se...

Embodiment 3

[0042] Example 3. gp120DNA vaccine construction

[0043] Plasmids pJW4303 and pGA4 / AE.gp120 (or pGA4 / BC.gp120, or pGA4 / ThB.gp120) were double digested with Nhe I and BamH I, respectively. The enzyme digestion reaction system is: 10×Buffer TangoTM 4 μL, 10×BSA 4 μL, plasmid 20 μL, NheI 1 μL, BamHI 1 μL, and water to 40 μL. After the digested product was subjected to 7g / L agarose gel electrophoresis, the target fragment of about 1400bp and the linearized pJW4303 were recovered and purified with a gel recovery kit (Agarose Gel DNA Purification Kit Ver.2.0, Dalian Bao Biological Co., Ltd.).

[0044] Place the agarose gel after electrophoresis under the ultraviolet detector, cut off the gel containing the target DNA, weigh the mass of the gel block with an analytical balance, and calculate the volume of the gel block according to 1 mg = 1 μL. Add 3 times the volume of DR-I Buffer, place in a 75°C water bath to heat and melt the gel block, mix well every 2 minutes until the gel blo...

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PUM

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Abstract

The invention belongs to the technical field of biological medicine, relating to an HIV-1gp120 gene consensus sequence optimized by codons and a gp120 nucleic acid vaccine. In the invention, a Chinese HIV-1 epidemic strain (A/E recombinant subtype, B/C recombinant subtype and ThB subtype) envelope protein gp120 consensus amino acid sequence is obtained by applying multi-sequence comparative analysis; a gp120 gene segment encoding the consensus amino acid sequence is prepared by using an artificially synthesized method, and the gene is optimized by the codons and combines preferences of mammalian cells and escherichia coli codons; and on the basis, HIV-1gp120 nucleic acid vaccine consisting of the gp120 gene and eukaryotic expression vector pJW4303 is constructed. The nucleic acid vaccine can be applied to animal and human immunization and expression and production of the gp120 protein.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and relates to a codon-optimized HIV-1 gp120 gene consensus sequence and a gp120 nucleic acid vaccine. Background technique [0002] Human immunodeficiency virus (human immunodeficiency virus, HIV) is an envelope (envelop, Env) virus, and the surface of each viral envelope is mosaic with trimeric spikes formed by glycoproteins gp120 and gp41. According to different Env, HIV-1 is divided into M, N and O three groups. The M group is the most prevalent in the world. According to the whole genome, it is further divided into 9 subtypes (A, B, C, D, F, G, H, J, K), and presents geographical regional distribution characteristics. Among them, subtype B is the most popular subtype (mainly found in North America and Europe), subtypes A and D are mainly prevalent in Africa, and subtype C is distributed in Africa and Asia. These subtypes form a representative HIV-1M group different clades on the phylog...

Claims

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Application Information

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IPC IPC(8): C07K14/16C12N15/49A61K48/00A61K39/00A61P31/18
Inventor 王世霞张明顺黄祖瑚卢山
Owner 王世霞
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