Quikchange Mutagensis

Question 1

What does quikchange mutagenesis require?

Answer 1

Suitable primers, template DNA, dNTPs and DNA polymerase eg. KOD

Question 2

What are the steps of quikchange mutagenesis?

Answer 2

Design suitable primers, place in a thermal cycler with dNTPs, DNA polymerase, template, buffer and cations. Perform 16-30 cycles of PCR. Treat with Dpn1 to digest the methylated parent strand.

Question 3

Describe the primers?

Answer 3

24-40 nucleotides long, complimentary to the template with one point mutation in the centre, 75oC melting temperature, no 5’ phosphate, 50% GC content.

Question 4

What is quikchange mutagenesis used for?

Answer 4

To introduce a desired point mutation into a DNA sequence.