Molecular genetic techniques

Question 1

What temperature is DNA denatured at in PCR?

Answer 1

94C

Question 2

What is the first process in PCR?

Answer 2

Denaturation

Question 3

Second process of PCR?

Answer 3

Polymerase

Question 4

Third process of PCR? Temp?

Answer 4

Primer 72c

Question 5

What charge does DNA hold and why?

Answer 5

Negative due to the Phosphate groups

Question 6

What does “Allele-specific mutation detection” distinguish?

Answer 6

Between two alleles that may only differ by one single nucleotide

Question 7

How do allele specific primers work?

Answer 7

The 3’ end is designed so that it base pairs with the changed nucleotide distinguishing between normal and mutant (assay will not work unless perfect)

Question 8

What is compound heterozygous?

Answer 8

Both alleles of a gene have mutations, but these mutations are different.

Question 9

When is PCR unsuitable or difficult to use (which mutations) ?

Answer 9

If gene is too big
Fragile X mutation
GC-rich regions hard to PCR

Question 10

Detection of unknown mutations uses what?

Answer 10

DNA sequencing

Question 11

In sequencing what causes the random termination of a strand and why?

Answer 11

Dideoxynucleotide - Has an H not and OH so cannot for a phopshodiester bond

Question 12

How do you read the sequence? what technique?

Answer 12

Gel electrophoresis and fluorescence