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
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[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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