requesting information on yeast promoter less vector pAS 201

Martin Hirst hirst at interomex.com
Wed Sep 24 22:52:44 EST 2003


Hi Franco,

While it is true that many of the original promoter bashing
experiments were done on plasmid borne promoter-reporter cassettes it
is clear that the chromosomal context of the reporter plays an
important role in gene regulation and should not be ignored.  A good
example of this effect can be seen with the MET3 promoter which is
tightly regulated in its native context (ie in its chromosomal
location) but becomes leaky when placed in a plasmid (either ARS-CEN
or 2 micron).  There are likely multiple factors that lead to this
affect, histone organisation differences, repressive flanking
sequences, disturbance of the promoter/regulator balance (2micron
plasmids present 50-100 copies of the promoter!!!) etc....


Cheers

Martin


On 9/23/03 9:30, "francov" <francov at cellbnt.anat.med.ualberta.ca> wrote:

> Hello Martin,
> could you please briefly explain what do you mean by assaying in chromosomal
> context?
> Thanks
> Franco.
> 
> 
>> ===== Original Message From Martin Hirst <hirst at interomex.com> =====
>> Hello Shara,
>> 
>> I guess the first question would be how are you planning on assaying the
>> promoter activity?  LacZ, GFP, nutritional marker....
>> 
>> If you already have an integrated reporter you might want to consider
>> recombining your promoter directly.  Alternatively you could integrate a
>> reporter directly behind your promoter.
>> 
>> I would argue that promoters should always be assayed in a chromosomal
>> context, preferably in their native positions.
>> 
>> 
>> Martin
>> 
>> 
>> On 9/18/03 20:00, "Sreihani at aol.com" <Sreihani at aol.com> wrote:
>> 
>>> Hi;
>>> 
>>> I am in search of a promoter less yeast vector for my research. The purpose
>>> is to put the promoter of my interest in a yeast vector and look at the
>>> results
>>> for my promoter activity. We were supposed to get a promoter less yeast
>>> vector by the name of pAS 201, but the company in charge was not able to
>>> provide
>>> the vector to us as promised. I have contacted the SGD at Stanford, and Dr.
>>> Nash
>>> has suggested that I post my request on your site for possible response.
>>> 
>>> Thank you in advance.
>>> 
>>> Shara
> 
> Franco J. Vizeacoumar,
> Ph.D. candidate,
> Dept. of Cell Biology,
> Faculty of Medicine and Dentistry,
> University of Alberta, Edmonton, Canada.
> Phone: 780 492 7407
> email: francov at cellbnt.ualberta.ca
> URL  : http://www.ualberta.ca/~francov
> 

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