Molecular Techniques And Diagnosis II

Question 0

Explain the importance of PCR in genetic testing

Answer 0

Can be used to amplify a small sample for analysis
Can investigate single base mutations
Can investigate small deletions or insertions

Question 1

Describe the molecular basis of DNA sequencing

Answer 1

Used to determine order if nucleotide bases
Sanger sequencing used ddNTP which terminate chain aa H at 3’
Use lots endonuclease to cut DNA at different places
Mixture of dNTP/ddNTP -> lots of different length chains
Use electrophoresis to work out sequence

Question 2

Describe methods of dna hybridaisation

Answer 2

Southern blotting - unlabelled DNA from hel electrophoresis is marked
Nylon is used to transfer fragments from electrophoresis
Hybridised with gene probe to label fragments

Northern - similar with RNA
Western - similar with protein

Question 3

Describe role of DNA hybridaisation in genetic testing

Answer 3

Investigate gene structure - deletion/duplication
Investigate gene expansion, triplet repeats
Investigate variation (DNA fingerprinting)

Question 4

How is PCR used in allele-specefic tests

Answer 4

Use primers specefic for sequence either side of allele of interest to amplify specefic allele

Use DNA probe that is complimentary to mutated allele

Question 5

How is restriction analysis used in allele specefic tests

Answer 5

Use one/multiple enzymes with restriction sites around the allele sk can analyse size of fragments

Question 6

Discuss the range of molecular techniques used to analyse DNA at the gene level

Answer 6

Restriction analysis
Gene cloning
Gel electrophoresis
Polymerase chain reaction
Southern blotting

Question 7

Discuss the range of molecular techniques used to analyse DNA at the chromosome level

Answer 7

Karyotyping - organaise according to chromosome number
Flourescent in situ hybridaisation
Detect mutations via allele specefic probes