7.1 Using gene sequencing

Question 1

Define genome

Answer 1

The total of all the genetic material in the organism

Question 2

Describe the process of PCR

Answer 2

1. DNA sample, DNA polymerase, primers, and a good supply of nucleotide bases are mixed together in a PCR vial
   Placed in a PCR machine
2. Heated to 90-95 for 30 seconds which causes the DNA strands to separate as the hydrogen bonds are broken down
3. mixture is then cooled to 50-55 to allow the primers to anneal to the single DNA strands
4. finally heated to 72 again for at least a minute. this is the optimum temperature for the DNA polymerase enzyme which builds up complementary strands of DNA

Question 3

What is DNA sequencing?

Answer 3

Identifying the base sequence of a DNA fragment

Question 4

How can DNA sequencing be used in medicine?

Answer 4

To screen for heritable conditions. The base sequence of a particular allele/gene can be compared to a person’s DNA to see if it is present (gene sequencing)

Question 5

How is PCR useful in forensic science?

Answer 5

Amplifying minute traces of DNA found at a crime scene/on a victim to give enough to develop a DNA profile (DNA profiling)

Question 6

Describe the process of DNA sequencing

Answer 6

1. DNA molecules chopped into fragments and replicated using PCR
2. Fragments separated into single strands
3. Further replication using a mixture of bases, including terminator bases with attached fluorescent tags that stop the process at a particular base
   - Separating the fragments allows the original sequence to be worked out

Question 7

What is the terminator base?

Answer 7

A modified version of one of the four nucleotide bases that halts the build up of a DNA chain once it is added

Question 8

Describe DNA profiling (fingerprinting)

Answer 8

Identification of repeating patterns in the non-coding regions of DNA
The pattern that restriction enzymes cut an individual’s DNA into fragments is unique, giving a pattern that can be compared with forensic samples and relatives

Question 9

How are introns used in DNA profiling?

Answer 9

Within introns there are short sequences of DNA that are repeated to form micro and mini satellites.
The same ones appear in the same positions on each pair of homologous chromosomes
The more closely related two people are, the more similarities in their DNA there will be

Question 10

Describe the process of southern blotting

Answer 10

1. An alkaline solution is added to the gel after electrophoresis which picks up the DNA fragments and forms ‘blots’ on the paper
2. The solution denatures the DNA fragments so the strands separate and the base sequences are exposed
3. Large numbers of gene probes are added to the filter which bind to complementary DNA strands in a process called hybridisation
4. Excess probes are washed away and the filter is placed under UV light to show up the DNA regions
5. A DNA profile is produced as a graph - each peak represents the number of microsatellite repeats in a fragment.

Question 11

What is a gene probe?

Answer 11

Short sequences of DNA that are complementary to specific sequences being sought