From Kanchen.Loganathan from WPAFB.AF.MIL Mon Jun 8 11:24:38 2009
From: Kanchen.Loganathan from WPAFB.AF.MIL (Loganathan,
Kanchen G CTR USAF AFMC 711 HPW/RHPB)
Date: Thu Jun 11 08:39:37 2009
Subject: [Yeast] lysis of yeast cells
Message-ID: <2B00361EE3107A4F88383EC1B041DC9A0567F63C@VFOHMLAO01.Enterprise.afmc.ds.af.mil>
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From she from bio.dtu.dk Tue Jun 16 06:02:31 2009
From: she from bio.dtu.dk (=?iso-8859-1?Q?S=F8ren_Helmark?=)
Date: Tue Jun 16 12:06:54 2009
Subject: [Yeast] Promoters for RNA pol I and III
Message-ID: <689B8BC5EB1B6A418FFE1D6997EAD881036BA7896A15@WINEXCHANGE1.win.dtu.dk>
Hi,
Do you know if promoters for RNA pol I and III are characterized and where I can find the exact sequence (databases/papers...)
Regards,
Soren
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From Jennifer.Chang from nyumc.org Thu Jun 18 11:17:58 2009
From: Jennifer.Chang from nyumc.org (Chang, Jennifer)
Date: Thu Jun 18 11:25:49 2009
Subject: [Yeast] No background with yeast two-hybrid screen
Message-ID: <628D3F39A80B5F47A4F231911D2B58CF01350DE0@MSGWSDCPMB02.nyumc.org>
Hi, All,
I was hoping that someone may have some insight into my problem as I am
quite stumped. I performed a yeast two-hybrid screen using Clontech's
new Matchmaker Gold system, which tests for interactions by growth on
the antibiotic Aureobasidin A (AbA, 200ng/ml concentration), appearance
of blue color with addition of X-alpha-Gal, growth on -HIS, and growth
on -Ade. My bait was in strain Y2HGold (which apparently is very much
like the strain in their Matchmaker 3 kit - AH109) and my library in
Y187. I thawed an aliquot of my library in Y187 (after thawing, the
titer was ~10^8 cfu/ml) and mated the library strain to the bait strain.
After mating the strains, I tested the viability of the diploids (~10^5
cfu/ml), the bait (~10^8 cfu/ml), and the library (~10^6 cfu/ml) on the
appropriate media and I have screened over 10^6 diploids as the company
manual says to. I initially tested for interactions on the lower
stringency media, which is -LEU, -TRP, X-alpha-Gal, AbA and there was
absolutely no growth. I retested for interactions on lower stringency
medium by reducing the amount of antibiotic to 50ng/ml, 100ng/ml, and
150ng/ml. I still have no growth even on the lowest concentration of
antibiotic. I then tested for interactions on -LEU, -TRP, -HIS,
X-alpha-Gal and I still get nothing! I am really surprised by this last
result as everything I have read suggests that HIS expression is very
leaky and needs to be suppressed by 3-AT in order to detect true
interactions.
I have previously tested to make sure that my bait does not
auto-activate, is not toxic, and is expressed. The positive and negative
controls provided by the company behave exactly as they should so I know
the media preparation is not the problem. Therefore, I am quite stumped
as to why I am getting absolutely no background, even on media (-HIS)
that is supposed to give high background. Any ideas on what could be
happening?
Has anyone worked with the Matchmaker Gold kit and what have your
experiences been?
Best,
Jennifer
Regards,
Jennifer Chang, PhD
Post Doctoral Fellow, Cell Biology
Medical Sciences Building, Room 698
New York University School of Medicine
550 First Avenue
New York, NY 10016
Tel 212-263-5316
Fax 212-263-8561
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From stephanie.kaut from uni-konstanz.de Tue Jun 23 14:45:25 2009
From: stephanie.kaut from uni-konstanz.de (Stephanie Kaut)
Date: Wed Jun 24 09:31:20 2009
Subject: [Yeast] yeast plates with oleic acid
In-Reply-To: <1263964104.20090623170659@js-kaut.de>
References: <1263964104.20090623170659@js-kaut.de>
Message-ID: <1ef12e50906231245m3c6377c3m29befa2c29bddadb@mail.gmail.com>
Hello together,
I want to test some yeast strains for growth on YPAD plates with oleic
acid.
But preparation of the plates seems to be tricky, because oleic acid and
the agar don't mix well, and the oleic acid swims on the top of the agar.
What can I do to have nice plates?
Best regards,
Stephanie
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From Bida.John from mayo.edu Wed Jun 24 15:11:49 2009
From: Bida.John from mayo.edu (John Bida)
Date: Fri Jun 26 08:34:16 2009
Subject: [Yeast] Promoters for RNA pol I and III
In-Reply-To: <689B8BC5EB1B6A418FFE1D6997EAD881036BA7896A15@WINEXCHANGE1.win.dtu.dk>
References: <689B8BC5EB1B6A418FFE1D6997EAD881036BA7896A15@WINEXCHANGE1.win.dtu.dk>
Message-ID:
I know the yeast three hybrid system uses the polIII promoter and
they have the sequence for the plasmid posted here:
http://www.biochem.wisc.edu/faculty/wickens/lab/3h.aspx
You can also search for non-coding RNA's here
http://www.yeastgenome.org/cgi-bin/search/featureSearch
and find polIII encoded genes and look at the papers that identified
them:
http://www.yeastgenome.org/cgi-bin/reference/reference.pl?
dbid=S000064699
~Jp
On Jun 16, 2009, at 6:02 AM, S?ren Helmark wrote:
> Hi,
>
> Do you know if promoters for RNA pol I and III are characterized
> and where I can find the exact sequence (databases/papers...)
>
> Regards,
> Soren
> _______________________________________________
> Yeast mailing list
> Yeast@net.bio.net
> http://www.bio.net/biomail/listinfo/yeast
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From franklymr.d from gmail.com Thu Jun 25 12:39:34 2009
From: franklymr.d from gmail.com (Frank Dixon)
Date: Fri Jun 26 08:34:21 2009
Subject: [Yeast] yeast plates with oleic acid
In-Reply-To: <1ef12e50906231245m3c6377c3m29befa2c29bddadb@mail.gmail.com>
References: <1263964104.20090623170659@js-kaut.de>
<1ef12e50906231245m3c6377c3m29befa2c29bddadb@mail.gmail.com>
Message-ID: <8ec00eeb0906251039q7bc36a68t953e4bae90ffef68@mail.gmail.com>
I have cultured yeast in wall board, and am running remediation tests.
If I need to isolate yeast species for Heat, cold envelope testing.
Or isolate for chemical testing or gel shipping purposes, do you
reccommend particular assay to separate cleanly from solid substrate
and identify? Currently we are using a modified already batched
saccharomeyes w/ecoli xylose breakdown courtesy of DoD operations here
in Santa Ana, Ca. You guys are the best! thanks Mr. D
On Tue, Jun 23, 2009 at 12:45 PM, Stephanie
Kaut wrote:
> Hello together,
>
> ?I want to test some yeast strains for growth on YPAD plates with oleic
> acid.
> ?But preparation of the plates seems to be tricky, because oleic acid and
> ?the agar don't mix well, and the oleic acid swims on the top of the agar.
> ?What can I do to have nice plates?
>
> Best regards,
> Stephanie
>
> _______________________________________________
> Yeast mailing list
> Yeast@net.bio.net
> http://www.bio.net/biomail/listinfo/yeast
>