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Insulin and Genetic Engineering

  • Diabetes mellitus is the inability of beta cells of pancreas to produce insulin
  • Restriction enzymes/endonuclease cut DNA at specific recognition sites
    • This produces either "sticky ends" or "blunt ends"
    • DNA ligase can be used to re-join the ends
  • Recombinant DNA technology combines the DNA from two different organisms
  • Reverse transcriptase catalyses the formation of DNA from mRNA
  • Vector is a gene carrier. It will carry a human gene into the cell of a bacterium or yeast that will be used to make human protein. Produces no benefit for viruses / carrier
  • Plasmid, circular strand of DNA, are useful vectors to make human protein from bacteria
  • Transgenic organisms contain another species DNA

Integration Link

[EXAM] Remove a particular gene from the DNA of an animal cell

  • Locate with the use of a gene probe
  • Use restriction enzymes
  • Use endonucleases to cut at specific base sequence by hydrolysing
  • Breaking sugar-phosphate bonds

[EXAM] Insert this gene into the genetic material of a bacterium

  • Same restriction enzymes
  • Cut at same base sequence in bacterial DNA
  • Leaving sticky ends/hydrogen bonds break
  • Join/splice with ligase
  • Use of plasmid

Task to find and insert the gene into bacterium for Insulin production

  • Isolate human gene, e.g. insulin, by using cytoplasmic mRNA (no introns)
  • Reverse transcriptase, taken from a retrovirus, makes DNA from mRNA
  • DNA is given "sticky ends" by using the enzyme restriction endonuclease
  • Insert into a plasmid from a bacterium
    • Dissolve cell walls using enzymes
    • Centrifuge to separate bacterial chromosome ring from plasmids
    • Cut open the plasmid
    • Add sticky ends
  • Mix plasmid and DNA gene together and use DNA ligase to stick them together
  • Mix with bacteria //only ≈1% will take up the engineered plasmids
  • Identify by using antibiotic resistance. Add gene for antibiotic resistance next to insulin gene in the plasmid. Add antibiotic to the culture / only bacteria surviving have insulin gene
  • Grow transformed cells using industrial fermenters
  • Isolate and purify human protein made by these cells

Moral and ethical issues associated with recombinant DNA technology

  • Transgenic bacteria or viruses may mutate and may become pathogenic
  • Genetically modified crops could "escape"
    • Forms a genetically modified population in the environment
    • Genetic modification may involve the resistance to herbicides
    • Escaped crops may become "superweeds" that are difficult to kill and control
  • Transgenic organisms could upset the balance of nature
    • Population of transgenic salmon have been produced in which individuals grow rapidly
    • These transgenic fish could compete for food with other fish species