• Genetic engineering involves the manipulation of naturally occurring processes and enzymes.
  • extraction/syntheses a gene(s) from one organism
  • transfer gene(s) into another organism (of same or different species)
  • gene is expressed in new host

 

Stages involved

  • Obtain a copy of the required gene
  • place the gene in a vector
  • introducing the vector DNA into the host cell
  • select the cells which have taken up the foreign DNA

 

Plasmids as vectors in gene cloning

  • Plasmids are small, circular pieces of double-stranded DNA
  • small ie they are easy to use
  • exist naturally in bacteria ie bacteria take up plasmids from surroundings
  • can be produced artificially
  • double stranded: can insert genes from prokaryotes and eukaryotes
  • replicate independently in bacteria
  • can be transferred between different bacterial species

 

Enzymes used

  • Restriction endonuclease:
  • restrict viral infections by recognizing and breaking down DNA of invading viruses.
  • binds to specific target site on DNA (sequence of bases) and cut sugar phosphate backbone
  • straight across produce blunt ends
  • staggered fashion produce sticky ends
  • sticky ends: short lengths of unpaired bases; easily form hydrogen bonds with complementary base sequences on other fragments of DNA cut with the same restriction enzyme
  • Reverse transcriptase:
  • uses single-stranded mRNA as the template
  • reverses the transcription process: produces single stranded DNA
  • single-stranded DNA + DNA polymerase = double-stranded complementary DNA (cDNA)

DNA ligase links together sugar-phosphate backbones of DNA molecules and plasmid produces a closed circle of double-stranded DNA containing the new gene ie recombinant DNA

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