Yeast two hybrid

Martin Hirst hirst at interomex.com
Thu Jun 6 18:57:56 EST 2002


Hi Trent,

Yes I have seen  multiple cDNA's rescued from a single two-hybrid positive.
This usually arises from yeast that are inadvertently carried over when you
pick your positives.  It can also be related to your prey copy number - but
I think the proquest system is low copy.  You can usually get around it by
re-streaking your positives back onto selective media and picking a single
colony from there.  pPC86 carries the AMP marker (PNAS 89, pp.5789-5793) - I
expect pPC97 does as well.


Cheers
Martin


Interomex Biopharmaceuticals Inc.
3590 West 41st Ave.
Vancouver, BC
Canada
V6N 3E6





On 6/4/02 12:06 PM, "Trent Bosma" <Trent_member at pathlink.com> wrote:

> Yeast two hybrid  (ProQuest system using ResGen Hela cDNA library)
> 
> 1.  The majority of my putative interactors contain more than 1 cDNA.  Has
> anyone experienced similar results and found a cure apart from changing
> my bait?  
> 
> 2.  Has anyone successfully extracted the plasmids out of the controls
> provided with the kit  i.e. what is the antibiotic marker of pPC97?.  I was
> expecting it to be kanamycin as with pPC86 but it doesn’t appear to be so.
> 
> Trent J. Bosma PhD.
> LCI/NIAID/NIH
> Bldg. 10/11N222
> 301 496 7680

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