Inserting Target Genes

Question 1

What are the 3 processes involved in inserting target genes?

Answer 1

isolating the target gene

inserting the target gene into a vector

inserting the vector into bacteria

Question 2

What happens during isolating the target gene?

Answer 2

• use restriction enzymes which will leave sticky ends e.g. insulin

Question 3

What happens during inserting the target gene into a vector?

Answer 3

a vector is something that moves DNA from one place to another place
e.g. virus - bacteriophage - virus that infects bacteria
plasmids - double stranded loops of DNA used to transfer genes between bacteria

• use the same restriction enzyme to cut the plasmid open
• the sticky ends will be complimentary
• DNA ligase will reform the phosphodiester bonds
• forms recombinant DNA - DNA from more than one source / organism e.g. plasmid and target gene

Question 4

What happens when you insert a vector into bacteria?

Answer 4

transformed organism - contains recombinant DNA

• Ice cold calcium chloride - heat shock them (stressing the bacteria and make the cell wall more
  permeable)
• or you can do electroportation (electrical current)
• both ways increase the permeability of the bacterial cell wall - more likely to take up the
  recombinant plasmid