Jordans Notes 7

Question 1

What region of DNA should a primer be designed for?

Answer 1

A conserved region

Question 2

What targets the same DNA in many different species for PCR?

Answer 2

Universal primers
can be broad (within family/order)
or narrow (within genus)

Question 3

What kind of solution is 1%agarose gel placed in for electrophoresis?

Answer 3

TBE Buffer

Question 4

How is fragment size determined in gel
electrophoresis?

Answer 4

run next to a DNA “ladder” which has fragments of standardized sizes

Question 5

What should you do with DNA if inhibitors prevent
PCR?

Answer 5

Dilute solution

Question 6

What to do if PCR returns bands of the wrong size

Answer 6

1. redesign primers if target band isn’t present
2. If you can see target band:
   a. increase T
   b. Increase primer length (specificity)
   c. touchdown PCR

Question 7

What does low heterozygosity indicate?

Answer 7

That most individuals in a population share the same alleles
Inbreeding or low genetic diversity

Question 8

What makes a ddNTP different from a dNTP?

Answer 8

ddNTPs don’t have a hydroxyl group at the 3 prime end of the sugar

aborts any further nucleotides binding to the 3’ position

Question 9

What two things do chaotropic salts do?

Answer 9

Destabilises hydrogen bonds
- denaturing proteins
Create a Hydrophobic environment
- makes silica membrane the most suitable binding partner for DNA

Question 10

What is difference between GBS and RADseq?

Answer 10

Shearing and size selection