In Vivo Cloning Flashcards

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1
Q

Outline of in vivo cloning

A
  • insertion of isolated gene into a vector (plasmid)
  • transformation: transfer of plasmid vector containing the isolated DNA fragment into a suitable host (bacteria)
  • identification of successfully transformed bacteria (contain the plasmid and DNA fragment)
  • replica plating
  • cloning of successful host cells
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2
Q

What is a gene marker?

A
  • used to identify which plasmids have taken up the DNA fragment
  • the gene marker is disrupted if the DNA fragment is present- the gene won’t be expressed
  • examples: antibiotic resistance, adding a fluorescent protein, making an enzyme with an identifiable product
  • if these are lost, the DNA fragment has been taken up (success)
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3
Q

Relplica plating

A
  • all bacteria grown on master plate
  • bacteria which grow on ampicillin but not tetracycline plate have been transformed successfully with plasmids
  • select these colonies from the master plate
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