Random Mutagenesis

[lcronan]

I am attempting to make a library of copy-number mutants. In order to do this, I wanted to use random mutagenesis to create the mutations. Does anyone know of a good, tried and true technique? I have tried a random mutagenesis kit, but so far have gotten no mutants. Of course, I may be doing something incorrectly, but I am not getting any helpful feedback from my boss... and yes, I have asked! If anyone has any advice, hints, knowledge, thoughts, etc. to share on the subject, I would be extremely grateful!

Thank you!
Leah

[mith]

what kit?

[lcronan]

Stratagene's Random Mut. Kit

[lcronan]

OK, I found a method that works:

Hydroxylamine Mutagenesis. I got sooo many mutants, now it's time for screening...

I tried one protocol that asked for incubation at a high temp for a short amount of time. This didn't make sense to me since hydroxylamine is denatured at high temps, but I tried it anyway and got no results. I tried another that asked for incubation at 37 for 20hrs and that seems to have worked beautifully!