human insulin case studies

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  • engineering case studies: human insulin
    • pig insulin
      • diadvantage: not identical to human insulin
      • disadvantage: only small amount can be obtained from each pig
    • small gene so difficult to locate.
    • centrifugation used to obtain mRNA for gene. reverse transcriptase used to make a copy producing a single strand of DNA
      • this is used as a template
    • free nucleotides added to template of DNA to produce double stand called cDNA
      • DNA polymerase
    • plasmids cut open with restriction enzyme and mixed with cDNA insulin gene
    • DNA ligase seals cDNA gene into cut plasmid
      • recombinant plasmids taken up by bacteria
    • some bacteria do not take up plasmid
    • some bacterial plasmids sealed back together and not recombinant
      • some bacteria do not take up plasmid
      • recombinant plasmids taken up by bacteria
    • if gene is present, bacteria will not be resistant to tetracycline but will be resistant to ampicillin
    • replica plating-the process of growing bacteria on an agar plate then transfering a replica of that growth to other plates, usually containinng different growth promoters or inhibitors
    • reverse transcriptase :retroviruses carry genetic material as RNA so when they infect a cell they cause reverse transcriptase to be synthesised in host cell.

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