lecture 19

Question 1

describe insulin structure

Answer 1

Insulin is a double-stranded molcule. With one alpha strand and one beta strand. They are help together by intermolecular disulphide bonds.

Question 2

why cant we possibly use pig and cow insulin in diabetic patients

Answer 2

But they have slight variation in a couple of amino acid residues. The difference between the human and animal versions is enough for the human body to reject pig insulin. Causing immune response.

Question 3

general description for DNA technologies

Answer 3

Joining bits of DNA together (sometimes from
different species). These are then inserted into a
organism to produce (express) a useful protein.

Question 4

what are plasmids

Answer 4

Plasmids are critical in biotechnology. The are usually Double stranded circular molecules in bacteria which are separate from the central chromosome, but they can be linear plasmids as well. Plasmids have their own replicative origins, this means they can replicate independantly of the main chromosome

Question 5

what are the key parts of a plasmid

Answer 5

Origin of replication- allows initiation of replication using host DNA polymerase.
Antibiotic resistance gene- this allows us to determine if the transformed gene was taken up by bacteria.
Promoter- drives gene expression in cells with the appropriate transcription factor machinery.

Question 6

where do restriction enzymes originate from and what it do?

Answer 6

Naturally found in bacteria – defense system to
degrade foreign DNA, Cut dsDNA at specific sequences

Question 7

how does ligase help dna recombinant technology

Answer 7

DNA ligase catalyses the formation of phosphodiester bond to repair nick in DNA backbone. this nick being made from the restriction enzymes

Question 8

sequence of events for amplifying plasmids and genes

Answer 8

Transformation = transfer of plasmids into
bacteria
* Transformed bacteria selected by
antibiotic resistance contained on
plasmid
* Expression of plasmid gene in bacteria (if
bacterial promoter).
* Amplification of bacteria and purification of
DNA for downstream uses e.g. PCR,
cloning, transfection into other cells or
organisms

Question 9

what do we have to remove from a gene before implanting into a plasmid and what do we do to get around this

Answer 9

Prokaryotic genes
* Don’t have introns
* Prokaryotes don’t have the machinery to
process eukaryotic introns
so we take mRNA and reverse transcribe it into cDNA and implant that into the cell