Genetic engineering

• Commonly used vector for bacteria?

  Plasmid
• A vector transports DNA fragments into a host cell
• How are DNA Fragments produced?

  Restricion endonucleoase is used to cut the desired DNA fragment into a fragment with sticky ends

  sticky ends on the DNA fragment are complimentary to the sticky strands on the plasmid as they should have been cut by the same restriction endonucleoase
• the role of DNA ligase during genetic engineering?

  DNA ligase is responsible for joining together the nucleotides of the DNA fragment with the nucleotides of the plasmid by catalysing the formation of phosphodiester bonds
• Explain why the same restriction endonuclease must be used to produce on both the DNA fragment and the plasmid during genetic engineering?

  The restriction endonuclease cuts both the DNA fragment and the plasmid at the same recognition sequence. This means that the sticky ends on both are complementary to each other.As a result, the DNA fragment and the plasmid can slot together with hydrogen bonds between the complementary bases
• Transformation is the process of getting the plasmid into the bacteria
• methods used in transformation:

  Electroporation - applying a small electrical current to make the cell membranes more porous

  Adding calcium ions to attract the plasmids closer to the cell membrane and then increasing temperature (heat shock) to increase fluidity of the membrane and allow it to pass through
• Issues during transformation in genetic engineering?

  • DNA fragments may join together
  • plasmid reconnects without DNA fragement
  • recombinant DNA does not enter bacteria
• What are marker genes?

  genes inputted into a bactera's plasmid that gives an observable characteristic that if the recombinant plasmid has been made successfully, will no longer show that characterisitic
• Example of marker genes used?

  Antibiotic resistance

  fluorescent genes

  enzyme producing
• benefits of using fluorescent marker genes over antibiotic resistance genes?

  No risk of antibiotic resistance being passed onto other bacteria

  Cheaper

  Observable

  Does not involve killing the bacteria that has the desired recombinant plasmid