M6 CH21

Question 1

Outline the steps in DNA profiling. (5)

Answer 1

1. extract DNA: PCR
2. digest sample: restriction endonucleases cut the DNA into fragments
3. separate DNA fragments: by gel electrophoresis
4. hybridisation: radioactive/fluorescent DNA probes added in excess to the DNA fragments on the membrane. they bind to complementary strands of DNA.
5. Observation: via either x rays or under UV light

Question 2

outline the steps involved in extracting the DNA for a DNA profile.

Answer 2

polymerase chain reaction:
1. temperature in the PCR machine is increased to 90/95c for 30secs, denatures the DNA by breaking the hydrogen bonds
2. the temperature is then decreased to 55/60c and the primers bind/anneal to the either end of the DNA strand
3. the temperature is increased again to 72/75*c for at least 1 minute. (this is the optimum temp for DNA polymerase to work) DNA polymerase adds bases to the primer, building up complementary DNA strands
result= 2x new DNA fragments

Question 3

explain how the DNA fragments are separated in DNA profiling.

Answer 3

by gel electrophoresis
molecules are separated according to their mass/charge
+ charged mols will move towards the cathode (- pole)
- charged mols will move towards the anode (+ pole)
tiny pores in the gel result in smaller molecules moving faster
- southern blotting: DNA fragments transferred from the gel onto a nylon membrane

Question 4

outline the uses of DNA profiling

Answer 4

forensics: creating DNA profile of blood samples found at crime scenes
analysis of disease risk: some non coding microsatellites have been found to be associated with increased risk of diseases. used to create risk assessments