D1.1 - DNA profiling (3t)

Question 1

what is the process of DNA profiling?

Answer 1

1. DNA samples are collected (from saliva, hair, blood or body cells)
2. PCR is used to amplify the DNA of particular markers, producing more copies
3. gel electrophoresis is used to separate DNA fragments according to their length, which is dependent on the number of repeats
4. gel electrophoresis generates a unique pattern of bands, which is called the DNA profile

Question 2

what is a marker?

Answer 2

a marker is a region of repetitive DNA sequences which varies in repeat number in individuals

Question 3

what are the uses of DNA profiling?

Answer 3

• forensic testing
• paternity testing
• detecting mutations
• diagnosing diseases by identifying the pathogen that has the person’s cells
• prepare DNA for use in biotechnological techniques
• force genetic changes through recombination PCR

Question 4

how is DNA testing used in paternity testing?

Answer 4

DNA sample is collected from both the child and possible parents and all the bands in the child’s profile must be in one of the parents’ profiles, because the child contains a combination of both parents’ alleles

Question 5

what is a PCR?

Answer 5

PCR is a polymerase chain reaction used to amplify DNA samples by making many copies

Question 6

what are the parts of a PCR test?

Answer 6

a PCR reaction mixture includes:
- sample of DNA to be amplified
- primers - short lengths of single stranded DNA with complementary bases which provide a starting point for Taq polymerase to synthesis a new strand of DNA
- free nucleotides to be used to create new DNA molecules
- Taq DNA polymerase which is used as it does not denature at high temperatures

Question 7

What are the 3 steps involved in PCR testing?

Answer 7

1. melting (denaturation) - mixture is heated to 90C, breaking the bonds between the two strands of DNA
2. annealing - temperature is lowered to 60C to allow the primers to bind to target DNA sequence
3. extending - optimum temperature for Taq polymerase (72C) allows it to form double-stranded DNA though complementary base pairing on a template strand

Question 8

why is cycling needed in PCR?

Answer 8

cycling is needed to make large amounts of DNA copies through constant cycling of low and high temperatures to promote melting and annealing of DNA strands

Question 9

why is PCR heated to a high temperature?

Answer 9

mixture is heated to a high temperature to separate DNA strands by breaking hydrogen bonds

Question 10

why is the temperature then lowered?

Answer 10

the temperature is then lowered to allow primers to bind to target DNA sequence

Question 11

how are new strands formed in PCR?

Answer 11

new strands are formed by complementary base pairing using Taq polymerase - used as it can withstand high temperatures and won’t be denatured

Question 12

what is gel electrophoresis?

Answer 12

gel electrophoresis is a tool for DNA fragment separation based on size

Question 13

what is the use of endonuclease restriction enzymes?

Answer 13

amplified DNA samples are fragmented using endonuclease restriction enzymes which cut DNA molecules at specific base sequences

Question 14

what is important to consider when comparing two examples?

Answer 14

when comparing two samples, they must be fragmented using the same endonuclease restriction enzyme

Question 15

how are DNA fragments separated?

Answer 15

DNA samples are separated by loading them into wells in an agarose gel and an electrical current is applied, causing the negative DNA to travel towards the positive anode

Question 16

what is the relationship between DNA fragment size and movement?

Answer 16

smaller DNA fragments move faster and further through the agaorse gel than larger fragments which are hindered by the gel matrix

Question 17

why must the gel be stained?

Answer 17

DNA must be stained to see the patterns of the bands produced