No Blue Colonies...I'm not that good.

Sergio seraln at _NO_SPAM_netscape.net
Tue Feb 19 04:49:55 EST 2002


I would say that the bacteria you're using is not alpha-complementable, it's 
just a lacZ- strain.
Vectors like pUC19 don't carry the complete LacZ gene, just a truncated form of 
it, and it only works within microorganism which carry the other part into their 
chromosome. If you need the blue/white selection, choose a strain with the 
phi-80 lacZ(delta)M15 mutation.

Hope this helps...
Sergio


JSUPolek wrote:

> I'm trying to shotgun clone in chromosomal fragments of a cellulolytic anaerobe
> into pUC19.  Everthing has gone well, and I'm using a Rec- strain of E. coli
> for transformation. But all I get are white colonies.  NO blue colonies at all,
> not even on the control bacteria with unligated plasmid.  Some plates had
> X-gal, and some had Bluo-gal.  Some had IPTG and some did not.  Amp
> concentration was 10mg/ml, and I did 1:1, 1:10, 1:100, and even 1:1000
> dilutions.  Colonies were in numbers in proportion to the dilutions, so they
> were my E. coli.  Why didn't any of them turn blue??  I don't necessarily need
> the b/w colony selection for what I'm screening for, but it would be helpful. 
> I would appreciate any help.  Thank you, S. Polek
> 




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