Method for preparing expressed gene for constructing apolipoprotein AIm of human

An apolipoprotein and protein technology, applied in recombinant DNA technology, botanical equipment and methods, biochemical equipment and methods, etc., can solve problems such as the inability to fundamentally treat cardiovascular and cerebrovascular diseases

Inactive Publication Date: 2009-07-22
CHENGDU CHEMPARTNER
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

In other words, drugs that inhibit arterial endogenous cholesterol production that are currently popular in European and American markets cannot fundamentally treat generalized atherosclerotic cardiovascular and cerebrovascular diseases.

Method used

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  • Method for preparing expressed gene for constructing apolipoprotein AIm of human
  • Method for preparing expressed gene for constructing apolipoprotein AIm of human
  • Method for preparing expressed gene for constructing apolipoprotein AIm of human

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Experimental program
Comparison scheme
Effect test

Embodiment

[0041] 1. Materials

[0042] 1. PCR-Kit was purchased from TaKaRa Company, restriction endonuclease, Taq enzyme, T 4 DNA ligase, T 4 Polynucleotide kinase and DNA molecular weight standards were purchased from TaKaRa Company, Promega Company and Jingmei Company, respectively.

[0043] 2. Strains and plasmids: BL21 (commercialized, purchased from the market), pET12a (purchased from Invitrogen) were preserved in our laboratory; pGEM-T was purchased from TaKaRa; pET12a / proApoAI m Built for this lab.

[0044] 3.PCR primers were synthesized by Shanghai Institute of Biochemical Cells.

[0045] 4. Reagents: Anti-human Apo A1 monoclonal antibody was purchased from Chemicon Interational Inc., Tris and SDS were purchased from AMRESCO, and Ni-NTA Agarose was purchased from Invitrogen.

[0046] 2. Method

[0047] 1. proApoAI m Preparation of gene DNA fragments and construction of engineering bacteria

[0048] (1) Extraction of total RNA from cells

[0049] i. Take 10 mg of human hep...

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Abstract

A process for preparing the expression gene used to configure human apolipoprotein AI milano includes using PCR primer to prepare pro-ApoAIm gene, configuring the recombinant plasmid of expression gene PET12a / PAPOAIm, configuring the engineered bacteria, fermenting, protein purifying of pApoAIm, and prepairng the dimer of pro-ApoAIm. It has high stability.

Description

technical field [0001] The present invention relates to the field of bioengineering, more specifically refers to the use of bioengineering technology to obtain human pApoAI m High expression methods of genes, including the synthesis of pApoAI m The design of PCR primer nucleotide sequence, the transformation method of gene mutation sequence, the construction method of recombinant engineering bacteria, the fermentation optimization expression method of engineering bacteria, pApoAI m Protein extraction method and pApoAI m Method of protein dimerization, and its medical use. Background technique [0002] Atherosclerotic cardiovascular and cerebrovascular diseases (such as coronary heart disease, stroke) are the main killers of fatal diseases in China and Western countries, mainly due to high serum total cholesterol (Gprdon et al.1981; Stamler et al.1986; Pooling Project Research Goup 1978; Anderson et al. 1987). Epidemiological survey revealed that for every 1% increase in...

Claims

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Application Information

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IPC IPC(8): C12N15/12C12N15/63C12P21/00
Inventor 龚邦强丁满生张梅芳
Owner CHENGDU CHEMPARTNER
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