Mutagenesis - (Jul/16/2001 )
I have cloned a receptor which contains a single base insertion causing a frame shift mutation. Due to time restraints and contamination problems it is not practicle to go back and redo PCRS.
Im trying to delete the inserted base using a Clonetech kit where you introduce a mutation to the receptor and one to the expression vector (pcDNA3) for slection purposes:
1. Has anyone used this method and if so does it have a high success rate?
2. My mutation is ~500bp away from the site that I will mutate in the vector - is this ok?
3. Any general advice about site directed mutagenesis in this setting?
Have you thought about the Quickchange kit form Stratagene, its worked every time in our lab.