Genomic library construction troubleshooting

[Duncan Clark]

Historians believe that in newspost 
<mailman.402.1236118891.13724.methods from net.bio.net> on Tue, 3 Mar 2009, 
Rosario Díaz-González <rdiazg from ipb.csic.es> penned the following literary 
masterpiece:
>Anything I'm missing? Any tip or hint?

What is the genomic and what is the E.coli.

You are using an mcr, mrr and hsdR minus E.coli? See appendix of NEB 
catalogue, just in case you are starting from highly methylated DNA.

I take it you have tested the following.

Cut vector with BamH I, no CIP, no ligase  -  should get back no 
colonies.

Now ligate and should get back thousands of colonies. All should have 
intact BamH I site.  Just proves that BamH I and ligase are working as 
they should etc.

Insert should self ligate - test by agarose gel - proves Sau3A I 
generates ligatable ends. Doesn't prove they are still GATC but it's 
step in the right direction.

Duncan
-- 
I love deadlines. I especially like the whooshing noise they make as
they go flying by.

Duncan Clark
GeneSys Ltd.