The subject of this should have been, "Homologous Recombination Again".
Basically this post is to sum up the responses people have emailed
me directly regarding my original post about homologous recombination.
My original post stated being a novice to Yeast Molecular Biology, is there
anything I should know about homologous recombination before I start my
experiments. More specifically, how does one prepare their fragments.
Replies were of the following nature:
1. Restrict your fragment out of the vector. It's better to gel purify
to make sure no contaminating uncut vector is around. It sounded like some
people just restrict and transform. I chose to gel purify.
2. Southern or PCR your (+) clones to make sure of intergration
That about sums it up. I actually transformed on 8/21 so colonies should greet
me tomorrow morning. In general thanks to all who replied, and specifically
there was one person I didn't personally reply back to (I can't remember
which response it was), thank you as well.
Dept. of Cell and Molecular Biology
5 Cummington St,
Boston, MA 02215
email: harkness at bio.bu.edu