in vivo

Question 1

vectors

Answer 1

a vector is a molecule that carries DNA into a host cell

• plasmids
• viruses
• liposomes

Question 2

plasmids

Answer 2

circular double stranded molecules present in some bacteria cells that have:

• one or more market genes eg code for resistance
• different restriction enzyme recognition sites

Question 3

viruses

Answer 3

• insert gene into the virus
• use this virus to infect the target cell - viral DNA is injected into the cell and gets replicated, transcribed and translated along with the cell’s own DNA

Question 4

liposomes

Answer 4

• phospholipid bilayer vesicles that contain DNA
• enter the cell by endocytosis

Question 5

problems with using viruses

Answer 5

• the gene may get inserted at any place within the host cell genome, may interrupt genes coding for essential functions - leads to cell death or cancers
• immune response against the virus in animals / humans - only low dosages can be used so lower efficiency

Question 6

insertion

Answer 6

DNA fragment placed in plasmid - vector - carries the DNA fragment into the host cell

restriction endonuclease - cuts out the DNA fragment and cuts the plasmid - complementary sticky ends which join together by H-bonds

DNA ligase - binds nucleotides on fragment and plasmid - catalyses formation of phosphodiester bonds

Question 7

what is formed

Answer 7

recombinant plasmid forms

Question 8

what needs to be added

Answer 8

promoter and terminator need to be added

Question 9

transformation

Answer 9

insert into bacteria - affected by cell membrane
due to plasmid being polar, hydrophilic and negatively charged

add calcium ions - positively charged - takes plasmid closer to membrane

heat shock - forms holes in membrane

applying high voltage

Question 10

what does the reaction require

Answer 10

ATP

Question 11

the 3 possibilities

Answer 11

• the plasmid re-circularises
• the insulin gene forms a circle
• the insulin gene is attached to the plasmid ie recombinant plasmid