HDTenEightyP/NetNews huggingface dataset:
From: djt2@po.CWRU.Edu (Dennis J. Templeton)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Genebank on CDROM?
Date: 30 Sep 92 18:23:22 GMT
Reply-To: djt2@po.CWRU.Edu (Dennis J. Templeton)
Organization: Case Western Reserve University, Cleveland, OH (USA)
NNTP-Posting-Host: thor.ins.cwru.edu


In a previous article, rrumpf@magnus.acs.ohio-state.edu (Robert Rumpf) says:

>I have heard that it is possible to get Genebank on a CDROM, and get updates 
>every so often...who do we call/write/email, how much is it, and does it work 
>on Macintosh CDROM readers?
>
>Bob
>
NCBI has announced a CDrom based genbank distribution for cheap.. $50-$60
 they have a package that is called entrez that contains the mac and pc 
search software for that price.  

Ask for a mailing from NCBI (202) 738-3238 (the fax number for orders 
is 202 512-2233)

Or contact Dennis Benson via Email (benson@ncbi.nlm.nih.gov)

NCBI also offers a retrieve service via email that is more up to date than
a cdrom could ever be.  It's a little clunky now, but improving.  For help,
Email retrieve@ncbi.nlm.nih.gov with the word <help> in the body of the message.

good luck,

dennis
Newsgroups: bionet.molbio.methds-reagnts
From: hsb@ucselx.sdsu.edu (Hirdeypal S. Bhathal)
Subject: ***Chicken cell line (third posting)****
Organization: San Diego State University Computing Services
Date: Wed, 7 Oct 1992 01:19:53 GMT




Hi netters

I am looking for an easy-to-grow, easy-to-maintain and easy-to-transfect
CHICKEN cell line. Just about any type of chicken cell line will meet our needs as long as it can be grown and transfected with relative ease. 
If anybody has such a chicken cell line I would appreciate hearing from you.
Any response is welcome. Thanks.


Hirdeypal Bhathal
Dept. of Chemistry
SDSU, San Diego 
CA-92182, USA
(619)594-3771
hsb@ucselx.sdsu.edu
hsb@chem.ucsd.edu
From: Klaus.Matthaei@anu.edu.au
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Sequence of Bluescript
Date: 30 Sep 92 01:27:07 GMT
Distribution: bionet

>This must be a FAQ, but I'll ask it once more: Is the sequence of the
>Bluescript (TM) cloning vector available somewhere? If not, does someone
>have an up-to-date restriction map? The Stratagene info sheet does not
>cover the "latest" enzymes. 
>
>-- 
>Martti Tolvanen, Dept. Biochem., Univ. Helsinki, Finland
>tolvanen@cc.helsinki.fi

G'Day
I have the following sequences on disk from Stratagene if anyone is
interested.

"On the enclosed disk are sequences from many of Stratagene's innovative
vectors.  There are 21 ASCII files contained on the disk, their names and
descriptions are as follows:


	Name	Description

	88B2KSM.DOC	pBluescript  II KS (-)
	88B2KSP.DOC	pBluescript  II KS (+)
	88B2SKM.DOC	pBluescript  II SK (-)
	88B2SKP.DOC	pBluescript  II SK (+)
	88BLKSM.DOC	pBluescript  KS (-)
	88BLKSP.DOC	pBluescript  KS (+)
	88BLSKM.DOC	pBluescript  SK (-)
	88BLSKP.DOC	pBluescript  SK (+)

	PBCKSM.DOC	pBC KS (-)
	PBCKSP.DOC	pBC KS (+)
	PBCSKM.DOC	pBC SK (-)
	PBCSKP.DOC	pBC SK (+)

	PBSPLUS.DOC	pBS (+)
	PBSMINUS.DOC	pBS (-)
	PBSPLSMD.DOC	pBS 

	PHSCPSK.DOC	Phagescript SK

	PMC1NEO.DOC	pMC1Neo
	PMC1PLA.DOC	pMC1Neo poly A

	PSG5.DOC	pSG5
	PXT1.DOC	pXT1
	PWE15.DOC	pWE15


The disk itself is preformatted for an IBM PC, and includes a copy of this
letter under contents.doc.

If you have further questions please contact Technical Services at
1-800-424-5444.


Sincerely,


Stratagene Cloning Systems"


Others are of course available from 'Vectors' in GeneBank.

Cheers, Klaus

--------------------------------------------------------------------------
Klaus Matthaei
Gene Targeting
The John Curtin School of Medical Research
The Australian National University
E-mail: Klaus.Matthaei@anu.edu.au

"If all else fails.  Read the instructions".
--------------------------------------------------------------------------
Newsgroups: bionet.molbio.methds-reagnts
From: zxmkr08@mailserv.zdv.uni-tuebingen.de (Cornelius Krasel)
Subject: Random mutagenesis by PCR?
Keywords: Manganese, PCR, mutagenesis
Organization: Comp. Center (ZDV) U of Tuebingen, FRG
Date: Fri, 2 Oct 1992 01:09:26 GMT

Dear Netters,

a colleague is searching for a method to mutagenize randomly parts of
a vector (i.e. the insert but not the necessary parts). In a recent
Science article (Science 257, 635: A.A.Beaudry and G.F.Joyce, Directed
Evolution of an RNA Enzyme) a method has been mentioned to do this
by using PCR. There are several references to journals which we are
completely unaware of:
Technique 1, 11(1989)
PCR Methods Appl. 2, 28(1992) (we have asked for a reprint of this
  one).

Maybe some of you has already used these or similar techniques (there
are rumours about adding Manganese to the buffer) and could enlighten
us? Please send email, I'll summarize.

Thanks in advance, Cornelius. 
--
/* Cornelius Krasel, Department of Physiological Chemistry, U Tuebingen    */ 
/* email: krasel@mailserv.zdv.uni-tuebingen.de (Internet)                  */
/*        krasel@chemie.uni-tuebingen.dbp.de (WIN/X400)                    */
/* "People are DNA's way of making more DNA." (anonymous)                  */
From: KNECHT@UCONNVM.UCONN.EDU
Newsgroups: bionet.molbio.methds-reagnts
Subject: (none)
Date: 2 Oct 92 18:12:55 GMT
Distribution: bionet

Hi Netters-  I remember reading somewhere that you can use ethidium bromide in
restriction digestions to make partial digests.  Does anyone know a protocol or
 reference to this technique.  Thanks- Dave Knecht (KNecht@uconnvm.edu)
Newsgroups: bionet.molbio.methds-reagnts
From: Z299CB@tamvm1.tamu.edu
Subject: Gas Chromatography Info or Reference Requested
Organization: Texas A&M University
Date: Wed, 7 Oct 1992 01:56:33 GMT

Need a general introduction to gas chromatography and mass spectroscopy.  Refer
ences would be appreciated!  Thanks.
 
Reply to: z299cb@tamvm1.tamu.edu
 
Dan Bergey
From: wetsel_r@wums.wustl.edu
Newsgroups: bionet.molbio.methds-reagnts
Subject: RE: hybridization problems
Date: 6 OCT 92 09:23:52 GMT
Organization: MIT PLASMA FUSION CENTER
NNTP-Posting-Host: msnews.wustl.edu

Klaus...  I publically and humbly apologize for changine your gender...
::grin::  so much for auto/reply & extract features...

David
From: tcritchl@sdcc3.ucsd.edu (Tod Critchlow)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Formaldehyde and DNA
Keywords: Is PCR Possible?
Date: 29 Sep 92 17:46:28 GMT


Hello,
I am looking for information on
how to repair DNA that has been fixed in formaldehyde.
In other words, is it possible to treat formaldehyde
fixed samples so that PCR is possible?

E-mail responses to me and I will post the
results.

thanks,
tod(tcritchl@ucsd.edu)
-- 
_____________________________________________________________________
Tod Critchlow(tcritchl@ucsd.edu)      |  What the Hell is a
Univ of Calif, San Diego              |  Signature anyways??
Scripps Institution of Oceanography   |
From: sbhattac@crc.ac.uk (Dr. S. Bhattacharya)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Anti-sense oligos
Date: 3 Oct 92 14:10:45 GMT
Distribution: bionet
Original-To: methods@uk.ac.daresbury

Please ignore this if my earlier e-mail got through...

Could anyone give me some guidelines on the design of anti-sense oligos
to target specific mRNA sequences in vivo (cell culture).  Is there a 
computer program that predicts "good" oligos?

Shoumo
From: BI1RGW@ibm.shef.ac.uk
Newsgroups: bionet.molbio.methds-reagnts
Subject: re: re: whence Coomassie
Date: 5 Oct 92 12:48:03 GMT
Distribution: bionet

Thanks to those who answered my query.... 2 more questionettes:

1. Given that some proteins w/o lysine lare stained by Coomassie, can anyone
furnish an explanation for the NON-staining of certain proteins?
An example is the D1 protein of the photosystem II reaction centre, except for
a few cases (such as in wheat) when (wouldn't you know it) there's an extra
lysine ;-)

2. How good is the linearity for conc:stain? Presumably this can only apply
for a particular protein, since different proteins seem to stain to differing
extents (or not at all).

Robin Walters                      Robert Hill Institute
                                   University of Sheffield
BI1RGW@IBM.SHEF.AC.UK              P.O. Box 594
                                   Sheffield S10 2UH.
+44 (0)742 768555 X4244            UK
From: wmelchior@NTBTOX.NCTR.FDA.GOV ("Bill Melchior, NCTR/FDA")
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Photographing Coomassie-stained Gels
Date: 29 Sep 92 16:26:41 GMT
Reply-To: wmelchior@NTDOC.NCTR.FDA.GOV
Distribution: bionet

>does anyone have the appropriate photographic settings/conditions to get
>good quality black&white 35mm shots of Coomassie Blue-stained PAGE gels?
>
>	- filters for contrast?
>	- regular B&W film (100/125 ASA) vs. high contrast?
>	- backlighting (lightbox) preferred?
>	- instructions for photoprocessing prints  
...
>we are familiar with the Kodak self-processing system but want negatives of 
>the gel as well.

MANY years ago I took photos of Coomassie gels, using Polaroid's P/N 55 film
(medium contrast) to get both prints and negatives; we already had an MP-4 
camera which took the film.  The only problem was the relatively slow speed 
(ISO 50).  If you actually want slides, Polaroid has slide holders.  We used 
an red-orange (Wratten 23A) filter for contrast.
________________________________________________________________________________
The opinions stated are mine, not those of NCTR or its sponsoring organizations.

Bill Melchior                                ||     OMNISCIENCE
National Center for Toxicological Research   ||    Knowing what
Jefferson, AR  72079                         ||    thou knowest not
(501) 543-7206                               ||    is in a sense
                                             ||    omniscience.
WMELCHIOR@NTDOC.NCTR.FDA.GOV                 ||       from Grooks, Piet Hein
From: naphade@ATLAS.CHEMISTRY.UAKRON.EDU (Madhavi J Naphade)
Newsgroups: bionet.molbio.methds-reagnts
Subject: (none)
Date: 5 Oct 92 22:48:49 GMT
Distribution: bionet

I would like to subscribe.
From: Klaus.Matthaei@anu.edu.au
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Hot Start PCR
Date: 6 Oct 92 01:32:47 GMT
Distribution: bionet

Re Hot Start

We use an alternative.  

We use a capillary cycler where the PCR is performed in a positive
displacement pipette tip. With an automatic Pipettor the PCR mix (say 15ul)
is loaded into the capillary containing everything but the target DNA
(RNA).  The target is then loaded in a separate volume (say 5ul) so the
primers and the target don't see each other until the temperature is raised
and the volumes mix by thermal convection.  Voila, hot start without
hassle.  A second benefit is that there is no oil overlay and therefore
less mess since the tubes are sealed by the plunger.  In our hands the
method is also much more efficient than conventional cyclers.

Cheers, Klaus
--------------------------------------------------------------------------
Klaus Matthaei
Gene Targeting
The John Curtin School of Medical Research
The Australian National University
E-mail: Klaus.Matthaei@anu.edu.au

"If all else fails : Read the instructions"
--------------------------------------------------------------------------
From: pnh@fcsparc6.ncifcrf.gov (Paul N Hengen)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: cm amplification?
Date: 1 Oct 92 15:15:49 GMT
Distribution: bionet
Organization: Frederick Cancer Research and Development Center

Tao Tao writes:

>I really don't think that you need to amplify pUC based vector, since each
>cell has about 200 copies.  What you need to do is to increase the final
>cell density, which could be achieved by better nutrition.  See BRL's
>focus or catalog about the TB (terrific broth) medium.

You don't have to use Terrific Broth to do this. Simply inoculate a
selective LB plate with 0.5 ml of log phase culture and incubate
24-36 hours. Then flood the plate with 3.0 ml of any kind of broth
or saline and scrape the cells from the surface with a glass rod.

Paul N. Hengen
National Cancer Institute
Frederick Cancer Research and Development Center
Frederick, Maryland 21702-1201 USA
From: djt2@po.CWRU.Edu (Dennis J. Templeton)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re:BclI and BamHI ligation
Date: 29 Sep 92 15:18:35 GMT
Reply-To: djt2@po.CWRU.Edu (Dennis J. Templeton)
Organization: Case Western Reserve University, Cleveland, OH (USA)
NNTP-Posting-Host: thor.ins.cwru.edu


Previously, on this channel...

>
>> Call me paranoid...but EVERYTIME I alkaline phosphatased a
>> vector, then gene-cleaned it, the ligations never worked.  NOTHING!
>> If I purify the fragment by gene-clean, then phosphatased it and
>> precipitated it by traditional means, the ligations always work!
>> I don't lnow why, but it appears that phosphatased, then gene-
>> cleaned vector doesn't work in my ligations.  Therefore I never
>> prepare my vector that way anymore.
>> 
>> Anne Howe
>> Glaxo
>> 

We have had good luck usin cip and home-made gene clean this way:

Do your digest, 37C, 60 minutes or so using 1 ug vector
Add 1ul cip (Boerhinger 1u/ul) for 5' ends or 5-10 u for blunts

37C for 15-20 minutes right in the RE buffer

Run the cut vector out on a gel, usually on the same gel as the insert prep

Cut out the band and recover with NaI/glass

ligate about 25-50 ng vector with a 10x excess of insert

This has two advantages: 

	1) if your digest is incomplete, you will both know it and remove
	   the uncut vector
	2) the gel + glass powder elution is good at killing the cip

And it's easy!

trust me.

p.s. to Anne at Glaxo... I'll trade tips for stock anyday.

dennis
Newsgroups: bionet.molbio.methds-reagnts
From: broe@aardvark.ucs.uoknor.edu (Bruce Roe)
Subject: Re: cycle seq. of lambda
Originator: news@midway.ecn.uoknor.edu
Date: Sat, 3 Oct 1992 13:04:00 GMT
Distribution: bionet
News-Software: VAX/VMS VNEWS 1.41    
Organization: University of Oklahoma - University Computing Services

In article <1992Oct2.132129.1956@gserv1.dl.ac.uk>, gessler@convex.HRZ.Uni-Marburg.DE (Gessler Manfred Dr.) writes...
>We would like to sequence lambda clones with cycle sequencing on an ABI
>sequencer. Has anyone done this with dye-terminators, or do we need labelled
>primers to get useful sequences? 
>Any tips on protocols for DNA prep, oligo length etc. are welcome!
> 
>Thanks in advance
> 
>Manfred Gessler
>Uni Marburg
> 
>gessler@convex.hrz.uni-marburg.de
>FAX (49)6421-285630

Manfred,
	We have tried this with labeled primers with mixed success.
The main problem was that template concentration needed to be optimized
and getting beautiful, intact template can be a chore.  Doug Berg with
the group at BRL and Palazzolo at LLB have done some studies. 

Try looking at:

 Phadnis, S.H., Huang, H.V. and Berg, D.E., Proc. Natl. Acad. Sci. USA 86,
 5908-5912 (1989).

 Strathmann, M., Hamilton, B.A., Mayeda, C.A., Simon, M.I., Meyerowitz,
 E.M.and Palazzolo, M.J., Proc. Natl. Acad. Sci. USA 88, 1247-1250 (1991).

for some ideas and suggestions (even these papers used radio-labeling).

Cheers.....bruce
  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
 \  Bruce A. Roe                     Dept. Chemistry and Biochemistry /
 /  BROE@aardvark.ucs.uoknor.edu     University of Oklahoma           \
  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
From: robison1@husc10.harvard.edu (Keith Robison)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Size of exons and introns?
Date: 1 Oct 92 16:25:32 GMT

naffara@crc.ac.uk (Dr. N.A. Affara) writes:

>This may have been asked before....

>Does anyone know the maximum, minimum, (if any) and average size of exons
>and introns in human DNA?

>The reason I am asking is that I am trying to design PCR primers from 
>cDNA sequence and I want to have some idea of the exon/intron sizes.

>Thanks,
>Phil
>(E-mail naffara@uk.ac.crc)


	I posted something a while back on the subject -- you might try
the archives.  The largest exon known is in apolipoprotein B and is
over 7 Kb.
	If memory serves me, human introns smaller than 75 bp are
very rare, but the largest ones can be enormous (10s of Kb, probably
some in the 100s of Kb).


Keith Robison
Harvard University
Department of Cellular & Developmental Biology
Department of Genetics / HHMI

robison@ribo.harvard.edu
From: sbhattac@crc.ac.uk (Dr. S. Bhattacharya)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Anti-sense oligos
Date: 3 Oct 92 00:11:44 GMT
Distribution: bionet
Original-To: methods@uk.ac.daresbury

Hi
Can anyone give me some guidelines on the design of 
anti-sense oligonucleotides for targeting specific
mRNA sequences in vivo (tissue culture). Is there perhaps
a computer program that does this?
Many thanks
Shoumo
From: rpgrant@vax.path.ox.ac.uk
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Looking for a good oligo
Date: 30 Sep 92 07:52:56 GMT
Distribution: bionet
Organization: Oxford University Molecular Biology Data Centre
Nntp-Posting-User: rpgrant

In article <1992Sep28.150413.3666@gserv1.dl.ac.uk>, sbhattac@crc.ac.uk (Dr. Shoumo Bhattacharya B1 061) writes:
> I have used the Primer program (from Eric Lander) with good results.  It
> is easy to use, and above all, is FREE.  I can recommend it.
> Shoumo

Ditto!!


Richard
From: jnsmar14@uctvax.uct.ac.za
Newsgroups: bionet.molbio.methds-reagnts
Subject: pH problem
Date: 4 Oct 92 16:58:26 +0200
Organization: University of Cape Town

I am having problems adjusting the pH of various 1 M solutions (citrate,
succinate etc.). My pH electrode, if left in these for a while (few minutes)
seems to drift, in the sense that when I rinse it with distilled water, and put
it back into my reference buffer, I get a lower pH value than I know it should
be. This obviously doesn't bode well for accurate pH's in the above mentioned  
solutions! What exactly is the influence of ionic strength on pH? I also don't
always get corresponding pH values with pH-paper. Can anyone tell me what is
happening to the electrode? 

Regards,
Pierre Janssens
Zoology Dept.
University of Cape Town
Newsgroups: bionet.molbio.methds-reagnts
From: vewart@kean.ucs.mun.ca
Subject: hybridization problems
Organization: Memorial University. St.John's Nfld, Canada
Date: Wed, 30 Sep 1992 14:12:21 GMT

To molecular biologists:
I am having a problem in probing Northern blot membranes ans slot blot 
membranes (Hybond N+). Sllight hazy background is evident on the 
membranes, suggesting that the probe is OK.  However, where the RNA is 
applied in slot blots or where it has migrated to in Northerns appears 
white.  The image looks like a negative.
	If you have encountered this problem or if you know of a 
possible solution, please leave a message here.
Thanks. 
Juan
From: dddzeij@cc.ruu.nl (van der Zeijst)
Newsgroups: bionet.molbio.methds-reagnts
Subject: microsatellites
Date: 29 Sep 92 18:24:50 GMT
Distribution: bionet
Original-To: methods@uk.ac.daresbury
X-Mailer: ELM [version 2.3 PL11]
















Who knows an easy way to detect the length variation in microsatellite fragments(CA repeats) without using radioactivity or expensive automatic sequence machines? Fragments are generated by PCR, are 60 to 100 bp long and contain a variable (CA)n (n=1

Hans Lenstra.
From: BI1RGW@ibm.shef.ac.uk
Newsgroups: bionet.molbio.methds-reagnts
Subject: whence Coomassie?
Date: 1 Oct 92 13:21:51 GMT
Distribution: bionet

Hello Netland.

Can anyone remind me of the details of what, how and when Coomassie R/G bind
to proteins?  As I recall, R binds slower but is more sensitive - but what do
they bind?  I have at the back of my mind some vague inkling about lysine
residues - but what about arginine, and what about the N-terminus?

I'm sure this is a FAQ, but...all comments, recollections, anecdotes welcome.

Thanks in advance,


Robin Walters                      Robert Hill Institute
                                   University of Sheffield
BI1RGW@IBM.SHEF.AC.UK              P.O. Box 594
                                   Sheffield S10 2UH.
+44 (0)742 768555 X4244            UK
Newsgroups: bionet.molbio.methds-reagnts
From: lhanson@moose.uvm.edu (lhanson)
Subject: Can anyone help with protein expression in pGEX vectors?
Organization: University of Vermont -- Division of EMBA Computer Facility
Date: Tue, 6 Oct 1992 00:39:39 GMT

	I've been trying to express 2 cDNAs in pGEX-2T.  Both cDNAs are
from a fungal library and encode proteins involved in mating reactions.
The predicted fusion protein should be about 67 kdal; glutathione S-transferase
(26 kdal) + cDNA encoded peptide (40 kdal).  I have followed the protocol
given in "Current Protocols in Molecular Biology" and haven't been having
much luck.  I can often see what appears to be an intensification of a
protein band around 70 kdal in induced cultures (as compared to an uninduced
control).  The fusion protein is expressed at significantly lower levels than
the parental vector alone expresses GST (gluathione S-transferase).

	Does anyone know of any "tricks" I might use to improve expression?
I've tried inducing in early log phase (1.5 hrs after inoculation) as well as
later (4-5 hrs post inoculation).  I've even tried growing plates of ~600-1000
colonies overnight and then suspending them in LB (as a slurry) and then adding
inducer (IPTG to 1-3.5 mM final).  This last technique works great for the
parental vector alone but not for fusions.  I think I've tried the other
obvious experiments such as upping the ampicillin to 200 ug/ml, checking
my plasmids to ensure they are the constructs I believe them to be, using
Terrific broth as opposed to LB.

	I'm after enough protein to use in rasing antibodies.  Affinity
purification of large cultures isn't the answer at this point either.  I
cannot get affinity purification of the small quantity of fusion protein
I can get expressed as I believe its in inclusion bodies!

	Lee Hanson
	lhanson@moose.uvm.edu

	Department of Botany
	University of Vermont
From: ED@micro.uct.ac.za (Ed Rybicki)
Newsgroups: bionet.molbio.methds-reagnts
Subject: faqs, old methods
Date: 2 Oct 92 10:36:27 GMT
Reply-To: ed@micro.uct.ac.za
Distribution: bionet
Original-To: methods@uk.ac.daresbury
X-Mailer: Pegasus Mail v2.1c R5.

Regarding FAQs and storage of useful tips that come up on the net:  I have
for some months been filing everything of a bacterial / viral / PCR nature
that has come up on methods-reagents (and on bionet.molbio-evolution) in a
random access database on our LAN in our Department, for general
access by our staff and students.  If anyone remembers, but cannot quite
place, a useful tip / method they saw some time back, send me a query and
I will send back whatever comes up on the subject from the database.  I
also collect references on PCR and on plant viruses if anyone is
interested.  The archiving system is worth a mention for the simple reason
thatr it is cheap and friendly, and can import almost anything, in any
format: it is ARCHIVIST, for PCs, from Oxford University Press, and is
probably obsolete by now as I think they have an update with a different
name.

Disclaimer: I don't represent anyone, I just use their products.
From: "michael coyne" <michael.coyne@channel1.com>
Newsgroups: bionet.molbio.methds-reagnts
Subject: cm amplification?
Date: 27 Sep 92 00:29:41 EST
Reply-To: "michael coyne" <michael.coyne@channel1.com>
Distribution: bionet
Organization: Channel 1 Communications


Fellow Netters:

Can anyone out there supply me with a protocol for chlorampenicol
amplification of pUC-based plasmids in E. coli?  I've seen various
non-specific references to it (ie in Maniatis), but all are written in a
conversational manner as kind of an aside, and do not provide specific
details.

I would greatly appreciate any information on a consistantly
reproducable method that someone out there's lab might use, including
details such as how much Cm to use, at what point in culture growth do
you add the antibiotic, etc.

Thanks -
Mike
michael.coyne@channel1.com
---
  SLMR 2.1a  Nothing is so smiple that it can't get screwed up.
--
Channel 1 (R)   Cambridge, MA