HDTenEightyP/NetNews huggingface dataset:
From: eepee@polaris.utu.fi (Esa-Pekka P{lvim{ki)
Subject: Re: What happens to 32P-labelled compounds?
Organization: University of Turku, Finland
Date: Thu, 30 Jul 92 08:58:12 GMT

In article <1992Jul27.201324.27564@access.usask.ca> goldie@herald.usask.ca (Hugh Goldie) writes:
>
>What happens to 32P-isotope labelled compounds when the 32P breaks down?
>I am interested in 32P-gamma-adenosine triphosphate, for example, and
>also oligomeric deoxynucleic acids labelled at their 5' termini with
>32P by the enzyme polynucleotide kinase.
>
>My old radioisotope text (Principals of Radioisotope Methodology, 3rd edn
>Chase G.D. and Rabinowitz, J.L., 1967.) indicates on page 151 that 32P decays
>to 32S by negatron emission and the table in the back of the book indicates 
>that 32S is a stable isotope.  Would the 32S atom merely replace the 32P
>atoms in these molecules?  Would the recoil of beta radiation damage
>adenosine triphosphate or oligonucleotides?

I understand that a S atom in ATP is replaced with a 32P atom to produce a
radioactive ATP molecule. When the 32P decays to 32S, the ATP molecule 
continues to exist as a "real" ATP with a 32S where it should be. 
I might be wrong, though.

/ep
From: mic268a@vaxc.cc.monash.edu.au
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re:PCR.PROB
Date: 26 Jul 92 14:17:09 +1000
Organization: Computer Centre, Monash University, Australia


>I have a colleague who is trying to PCR single
> copy genes from eukaryotic DNA by both normal
> and inverse PCR procedures. 30 cycles usually
> results in faint or non-existent bands being
>present at the appropriate sizes. However, when
> these bands are excised and re-PCR'd, then
>numerous smaller fragments are generated, half
>the size and smaller than the original band. Does
>anyone know why, and/or what can be done to prevent it.

>Thanks in advance,

>Mike Poidinger
>Dept of Microbiology
>University of Reading


Mike,
I have also been attempting similar experiments to your colleague.
I am using inverse PCR to amplify a flanking region to a transposon insertion
Initially I could only see a faint band of the expected size. When I re-PCRed
the PCR fragment I predominantly got a very small fragment and I did not get
the expected fragment.
What I then did was I increased the volume of the ligation from 20ul to 500ul
then I EtoH precipitated the ligation mixture and did the PCR on that.
To my surprise I got the expected fragment and got buckets of the stuff.
My suggestion to your colleague is to increase the volume of the ligation. Your
conditions are probably right, I would certainly not increase the number of
cycles. Hope this is of some help

Mike L.
Monash University
Clayton, Victoria
AUSTRALIA
From: shibumi@turbo.bio.net (Kenton A. Hoover)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Is this newgroup archived anywhere?
Date: 27 Jul 92 20:45:37 GMT
Reply-To: shibumi@turbo.bio.net
Organization: GenBank Computing Resource for Mol. Biology

gilbertd@sunflower.bio.indiana.edu (Don Gilbert) writes:
>ftp.bio.indiana.edu:/usenet/bionet/...
>contains this and other bionet groups back to december 1991.

We also have a complete set of bionet newsgroups on genbank.bio.net, which
is usually available (but not at this moment -- fixing something) via the
biosci waisserver on genbank.bio.net.


| Kenton A. Hoover                                 |                          |
| BIOSCI Network Administrator (bionet newsgroups) |    shibumi@presto.ig.com |
| GenBank/IntelliGenetics, Inc.       415 962 7300 |  shibumi@genbank.bio.net |
|=============================================================================|
|   This is the allegory of an individual christened Jed. An unprosperous     |
|   ascetical recluse who just about sustained his clan. Then it came to      |
|   pass one morning. He was in quest of nutriments, when elevated from the   |
|   soil arose some effervescent raw petroleum products.                      |
|                                                                             |
|   The fundamental issue one recognizes, venerable Jed's become excessively  |
|   affluent.  His indigenous relatives asserted, "Jed, depart from your      |
|   current domicile".  They pronounced California as the locale of           |
|   choice, so the household loaded up their property and possessions and     |
|   seceded to Beverly Hills.                                                 |
From: scottk@nuclease.berkeley.edu (Number Ten Ox)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Smiles on Southerns
Keywords: Smiles
Date: 30 Jul 92 10:52:24 GMT
Distribution: bionet
Organization: Biochemistry and Molecular Biology, U.C. Berkeley
NNTP-Posting-Host: nuclease.berkeley.edu

In article <3653@fcs280s.ncifcrf.gov>, pnh@fcs260c2.ncifcrf.gov 
(Paul N Hengen) writes:

|> -> Problems can occur when there is overloading with DNA, the
|> concentration of agarose is too high, or you've pushed it so
|> that the gel runs faster than normal. This causes a trailing
|> effect, mostly due to shearing of the DNA. If you're using very
                         ^^^^^^^^^^^^^^^^^^^

Huh? Can you explain how shearing the DNA causes it to *trail*
on a gel?


-- 
       ____         Scott Keeney, DNA repair-queer         ____
       \  /           scottk@mendel.berkeley.edu           \  /
        \/        Biochemistry and Molecular Biology        \/ 
                            U.C. Berkeley
From: pnh@fcs260c2.ncifcrf.gov (Paul N Hengen)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Smiles on Southerns
Keywords: Smiles
Date: 29 Jul 92 15:13:52 GMT
Distribution: bionet
Organization: Frederick Cancer Research and Development Center

In article <1992Jul28.110853.24275@gserv1.dl.ac.uk> MORRIS@ch.unige.cmu
("Mike Morris, Geneva") writes:

>Why do bands smile (not across the gel, but within each band) on 
>genomic Southerns?

I've corrected the problem in several ways:

1. I hardly ever run a Tris-acetate gel anymore. The Tris-borate gels
produce a much cleaner band (in my hands).

2. I never use glycerol in the loading buffer. Ficoll or perhaps
sucrose is better.

3. I start off the gel run with a very low current for about 5 to 10
minutes until the dye front is most of the way into the gel. This
allows the DNA to uniformly enter the gel at the side of the well.

4. I always stain with Ethidium Bromide AFTER the gel run.

-> Problems can occur when there is overloading with DNA, the
concentration of agarose is too high, or you've pushed it so
that the gel runs faster than normal. This causes a trailing
effect, mostly due to shearing of the DNA. If you're using very
large fragments of DNA for Southerns, you may have to run the
gel very slowly and keep the concentration of agarose to about
0.8 %. Also, make sure the buffer within the gel is the same as
that in the gel box. 

Good Luck. -Paul.
From: nwhite@uk.ac.crc (Mr. N.J. White)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Automated DNA extraction
Date: 21 Jul 92 17:30:32 GMT
Distribution: bionet
Original-To: methods@uk.ac.daresbury

Is there anyone out there with experience of automaed DNA extractors?

I have several hundred frozen samples (mainly leucocytes in a lithium
acetate/lithium dodecyl sulphate lysis buffer) from which I need to
extract DNA. I am looking at the possibility of using an automated
procedure but would like some opinions of the various systems before
I spend such a huge chunk of grant.

Please mail me at the address below with any suggestions and ideas.

Thanks

Nick White			e-mail:njw@mrc-lmba.cam.ac.uk
Haem. Med.			tel   :0223 336835
Cambridge University
MRC Centre
Hills Road
Cambs CB2 2SG
Newsgroups: bionet.molbio.methds-reagnts
From: cup@bones.biochem.ualberta.ca (Chris Upton)
Subject: Re: SDS in Flakes
Reply-To: cup@bones.biochem.ualberta.ca
Organization: University of Alberta
Date: Thu, 23 Jul 1992 14:31:01 GMT

I have some "super-pure" SDS from Millipore for 2D protein gels 
that comes as small flakes. I don't know about their regular SDS.

Cheers,
 Chris

---
Chris_Upton@darwin.biochem.ualberta.ca
Biochemistry Department
University of Alberta
Edmonton, AB T6H 2H7
Canada

Newsgroups: bionet.molbio.methds-reagnts
From: agoodrid@vaxa.weeg.uiowa.edu (Stephen Klautky)
Subject: Taq errors during PCR
News-Software: VAX/VMS VNEWS 1.41    
Organization: University of Iowa, Iowa City, IA, USA
Date: Wed, 29 Jul 1992 02:28:00 GMT

I have read a few papers indicating that Taq polymerase will
occasionally (about 1/500) misincorporate a base.  So far, the
impression I get is that these are random events.  If one sequences
the PCR product the misincorporations are not detectable.

Does anyone know if misincorporations can be consistent?  If this was
a problem it would be detected by sequencing the PCR product.


-=] Steve [=-      AGOODRID@VAXA.WEEG.UIOWA.EDU

Newsgroups: bionet.molbio.methds-reagnts
From: horton@molbio.cbs.umn.edu (Robert Horton)
Subject: Re: ethanol precipitation
Organization: University of Minnesota
Distribution: bionet
Date: Fri, 24 Jul 1992 19:05:29 GMT

I add the salt to the EtOh: 2% (w/v) KOAc in 95% EtOH.
-- 
Bob Horton            /\ "Crash programs fail because of the theory that
U. of Minnesota, CBS  || with nine women pregnant you get a baby a month" 
1479 Gortner Ave.    /||\   -Werner von Braun.  Disclaimer:"Bob who?"
St. Paul, MN 55108    ^^   horton@molbio.cbs.umn.edu/(612) 624-3790
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Cloning PCR products?
From: cytogen@chmeds.ac.nz (Martin Kennedy)
Date: 27 Jul 92 09:54:53 +1300

In article <d1gmx3m@lynx.unm.edu>, kim@m44.unm.edu writes:
> In article <1992Jul23.030739.10401@menudo.uh.edu>, bchs1b@Elroy.UH.EDU (Michael Benedik) writes:
>>
>>	There are a lot of different methods around for cloning PCR 
>>products to get around the problems of the fragment ends left after 
>>amplification. Does any one know whether this is solely a problem with
>>TAQ polymerase? Are there other polymerases which leave nice ends
>>suitable for subsequent cloning without going through the standard
>>gyrations?
> 

Apparently the use of thermostable polymerases with 3' to 5' exonuclease
activities (such as Vent) gives products with flush ends, as the nuclease
removes the troublesome 3' nucleotides  (NAR 20, p 144).

Cheers,   Martin
Newsgroups: bionet.molbio.methds-reagnts
From: rrumpf@magnus.acs.ohio-state.edu (Robert Rumpf)
Subject: Sequenceing Gel Fragment Aligning Software?
Organization: The Ohio State University
Date: Tue, 21 Jul 1992 23:12:24 GMT

I am looking for a Macintosh program which will allow entry of DNA sequences 
(say, off of a sequencing gel) and then do a simple alignment to allow me to 
string together contiguous readings/gels.  Currently I'm using DNAid+ 1.0, 
which does the job just fine except it crashes 90% of the time before even 
opening a document.  An updated version of this would be great, or clues as to 
how to make it run consistently.

Bob
Tue, 28 Jul 1992 12: 10:28 +0100
               Tue, 28 Jul 1992 12: 08:46 +0100
               Tue, 28 Jul 1992 14: 08:00 +0100
               Tue, 28 Jul 1992 14: 08:00 +0100
               Tue, 28 Jul 1992 14: 08:00 +0100
From: MORRIS@ch.unige.cmu ("Mike Morris, Geneva")
Newsgroups: bionet.molbio.methds-reagnts
Subject: Smiles on Southerns
Date: 28 Jul 92 13:08:00 GMT
Distribution: bionet
X-Envelope-To: methods@daresbury.ac.uk
Content-Identifier: Smiles on Sou...
X-Vms-Cc: MORRIS
Original-To: methods@uk.ac.daresbury
X-Vms-To: IN%"methods@daresbury.ac.uk"

Hi netters

Why do bands smile (not across the gel, but within each band) on 
genomic Southerns?
A few years back, I switched from ficoll to sucrose loading dyes,
cos conventional wisdom said the problem would go away. CW was right,
but the problem has recently and suddenly returned.
The smiles are worst when there is a lot of DNA (or perhaps salt in
the DNA sample?) ina a lane. 
Explanations and tips appreciated!

Mike Morris, Geneva, Switzerland
email morris@cmu.unige.ch
From: rybicki@uctvax.uct.ac.za
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: ethanol precipitation
Date: 21 Jul 92 11:29:37 GMT
Distribution: bionet
Organization: University of Cape Town

In article <920716092147.20200245@ntet.nctr.fda.gov>, WMELCHIOR@NTET.NCTR.FDA.GOV (Bill Melchior, NCTR/FDA) writes:
>>The problem I have with the use of ammonium acetate is the
>>requirement for the addition of two volumes of salt in addition to
>>the ethanol.  This usually screws up the maximum volume that I can
>>handle in a 1.5 ml tube.  Any suggestions for avoiding this, like
>>using higher concentration of amm. acetate?
> 
> This can be a problem.  My procedure is to add 1/2 volume of 7.5 M
> AmOAc, followed by twice the NEW volume of EtOH, giving a final volume
> 4 1/2 times as great as that of the original DNA solution.  If anyone has
> had experience using an even higher salt concentration, I too would like 
> to hear it.

1/5th volume of 10M amm acetate, 1 or 1/2 vol isopropanol does it quite well -
and is volatile, and works for sequencing PCR products too....
 _________________________________________________________________________
|                                 |                                       |
|  Ed Rybicki                     |    Now you've got the hang of it      |
|  Dept Microbiology              | There's nothing you can't do with it  |
|  University of Cape Town        |         If you're very into it        |
|  PB Rondebosch 7700             |         You can't go wrong....        |
|  South Africa                   |                                       |
|  fax 27 21 650 4023             |              - Mad John               |
|  ed@micro.uct.ac.za             | (Ogden's Nut-Gone Flake, Small Faces) |
|_________________________________|_______________________________________|
Newsgroups: bionet.molbio.methds-reagnts
From: txt15@po.CWRU.Edu (Tao Tao)
Subject: Question on Primer design for Cross-over PCR...
Organization: Case Western Reserve University, Cleveland, Ohio, (USA)
Date: Fri, 24 Jul 92 19:35:03 GMT


I would like to get fellow netters opinion on the design of primers for
cross-voer PCR.  Specifically, what's the minimum length of overlap? Is
15 bp long enough (with Tm ~45 degree)? 

Another question related to this is, what is the "ideal" procedures to
generate the hybrid molecule for next step cross-over PCR?  Is it required
or prefered that the two product be made using asymmetric PCR?

Thanks in advance for your reply.

Tao
From: broe@aardvark.ucs.uoknor.edu (Bruce Roe)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Buffer question
Date: 26 Jul 92 12:58:00 GMT
Organization: University of Oklahoma - University Computing Services
News-Software: VAX/VMS VNEWS 1.41

In article <14mn4tINNim9@matt.ksu.ksu.edu>, rdna@matt.ksu.ksu.edu (Paul Paukstelis) writes...
> 
>What is TBE, where can it be ordered, and are there any viable substitutes that
>can be used?  Could EDTA possibly be one of these?
>I'm a senior in high school doing a research project. Funds are short and TBE
>is getting hard to find.  Ohh yeah, what precent solution should the TBE in.
>Thanks a bundle in advance
> 
>Paul Paukstelis
>industriaL musiC froM manhattaN kS
>rdna@matt.ksu.ksu.edu

Paul and others:

	A number of years ago we switched to "modified TBE" which
has double the Tris and half the borate as originally described for
polyacrylamide gels.  Here's the composition of a 10-times concentrated
stock solution.  BTW, mTBE gives "cleaner" gels my increasing the
buffering capacity and has a slightly different pH than TBE.

10x Modified Tris-EDTA-borate buffer (mTEB; for polyacrylamide gels): 
	162 g. of Tris base, 27.5 g. of Boric acid, 9.3 g. of EDTA, 
	distilled water to 1 liter.

  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
 \  Bruce A. Roe                     Dept. Chemistry and Biochemistry /
 /  BROE@aardvark.ucs.uoknor.edu     University of Oklahoma           \
  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
Newsgroups: bionet.molbio.methds-reagnts
From: broe@aardvark.ucs.uoknor.edu (Bruce Roe)
Subject: Re: E.coli genotypes
News-Software: VAX/VMS VNEWS 1.41    
Organization: University of Oklahoma - University Computing Services
Distribution: bionet
Date: Sun, 26 Jul 1992 13:10:00 GMT

In article <29036.9207240840@crc.ac.uk>, sbhattac@mrc-crc.ac.uk (Dr. Shoumo Bhattacharya B1 061) writes...
>Many thanks to all those who responded to my query on E.coli
>genotypes.  To summarise, the info can be had from company catalogues, ATCC
>and Barbara Bachmann at Yale.  I wish someone would do an online database
>on this though.  Perhaps Genbank?  Then we could put in a request for 
>e.g. recA1 + dam-  +  whatever you please and see if any such
>bugs are available.
>Shoumo Bhattacharya
>..
	The ATCC does have dial-up access to their database but
it is not yet on the InterNet.  This info was given in their latest
NewLetter, and I don't have the details handy.  Maybe someone else
does.
  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
 \  Bruce A. Roe                     Dept. Chemistry and Biochemistry /
 /  BROE@aardvark.ucs.uoknor.edu     University of Oklahoma           \
  - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
From: djt2@po.CWRU.Edu (Dennis J. Templeton)
Newsgroups: bionet.molbio.methds-reagnts
Subject: SS Phagemid gradients, KI or CsCl?
Date: 21 Jul 92 15:40:49 GMT
Reply-To: djt2@po.CWRU.Edu (Dennis J. Templeton)
Organization: Case Western Reserve University, Cleveland, OH (USA)


We're trying to purify a mg or so of phagemid DNA, and need it to be free
from contaminating double stranded DNA, such as might come from broken
cells.  The protocol (sketchy) we're following suggests using EtBr/KI
gradients for this, and collecting the lower band (SS) and discarding the
upper band.  Our first go at this was a bust, as we lost almost all of the
DNA in the work up. (the KI solution containing the DNA was sucked up into
the H2O-saturated N Butanol!)

I measured the density of the KI solution (1.6) and it is the same as our
CsCl solution.  Obviously DS DNA floats in both near the middle of the
tube.

It made me ask myself, why KI and not CsCl?  Also, since we're pulling the
lower band (we see only two, the lower is brighter) mightn't it be
contaminated with some superhelical DNA along with the SS DNA?

I really hate not knowing what's behind the reasoning for this protocol.

Insights for why KI is used here and not CsCl would be appreciated, as
would any favorite methods for purifying SS DNA away from DS DNA.  Some
have suggested a low melt gel, but I'm hesitant, since we need a minimum of
50 ug for each run.

thanks 

dennis
Newsgroups: bionet.molbio.methds-reagnts
From: bchs1b@Elroy.UH.EDU (Michael Benedik)
Subject: Cloning PCR products?
Reply-To: bchs1b@Elroy.UH.EDU
Organization: University of Houston
Date: Thu, 23 Jul 1992 03:07:39 GMT


	There are a lot of different methods around for cloning PCR 
products to get around the problems of the fragment ends left after 
amplification. Does any one know whether this is solely a problem with
TAQ polymerase? Are there other polymerases which leave nice ends
suitable for subsequent cloning without going through the standard
gyrations?
	---------------------------------------------------
	 Michael Benedik
	 Department of Biochemical and Biophysical Sciences
	 University of Houston
	 INTERNET: Benedik@UH.EDU	BITNET: Benedik@UHOU
Newsgroups: bionet.molbio.methds-reagnts
From: gmc@helix.nih.gov (Gordon M. Charrick)
Subject: Is this newgroup archived anywhere?
Organization: National Institutes of Health, Bethesda
Date: Mon, 27 Jul 1992 13:44:22 GMT

I had a request from a scientist here at NIH looking to search through
postings that had already expired. Is there an archive site for this and
any other bionet newsgroups?

If someone is going to post the information, could he/she also send it to
me via email as I don't read this group.


                                       Thanks in advance,

                                            Gordon
--
Gordon Charrick [n4ytx]                (w): 301-496-4823
gmc@helix.nih.gov
Nat'l Inst. of Health, Bethesda, MD
Newsgroups: bionet.molbio.methds-reagnts
From: dresnick@athena.mit.edu (David I Resnick)
Subject: Re: S35- met labelling of proteins: URGENT
Organization: Massachusetts Institute of Technology
Date: Fri, 31 Jul 1992 00:00:28 GMT

In article <13702@gazette.bcm.tmc.edu> ak990140@SPECIALK.IAIMS.BCM.TMC.EDU (Arthur Kania) writes:

>  I'm trying to label proteins secreted by cultured cells. I would
>  like to know how much labelled amino acid I should add in order to
>  label adequate amounts. The medium is already met-deficient.
 
It depends on the cell line/how heavily your protein expressed/how
many methionines your protein has.  We do the following for proteins
secreted from CHO cells.  Note that if you want to label for very long
periods (overnight), you may have to add a bit of cold methionine as
well, or your cells will actually consume all of your 35S-met, and
stop producing protein.

1) Day 0:
   Set cells @1.5x10^5 per well in 6 well dishes

2) Day 2:
   Wash cells 2xPBS (to remove free methionine!)
   Add labelling media
       Hams F12-met
       80 uCi/ml 35S-Met or 35S-Express mix
       5% DIALYZED FBS (dialyze vs 150 mM NaCl, sterile filter)
	  You can leave this out.  The cells are perfectly happy
	  without serum for reasonable periods of time
   Incubate 5 hr, harvest media, spin at 1500xg to remove floating cells.

Notes and thoughts
      The quantity of 35Smet that you use and the length of time that
you label for obviously will be determined by the level of expression
of your protein.  We have been using stuff from New England Nuclear
called 35Express (or something like that).  It is a hydrolysate of
proteins from bacteria which have been fed 35SO4.  It is relatively
cheap - $99 for 5 mCi.


>  Thanks for Your help.

No problem.


--
       David Resnick                           dresnick@athena.mit.edu
From: themmen@nl.eur.fgg.endov ("Axel P.N. Themmen")
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: Tracer dyes in agarose gels
Date: 29 Jul 92 16:19:32 GMT
Reply-To: themmen@nl.eur.fgg.endov
Distribution: bionet


 > Hello. For some time I've been using a ficoll based loading
 > solution for my agarose gels with both xylene cyanol and
 > bromphenol blue added as tracer dyes. Recently, I saw an
 > advertisement for a loading dye mix of three dyes used.
 > When I called the company which sells these, they would not
 > tell me what the dyes are. BUT, they told me that they are
 > NOT the two mentioned above...So, I ask if anyone knows of
 > any other dyes that can be used for this purpose? In particular
 > I'm wondering if anyone uses a dye that migrates faster than the
 > bromphenol blue front?
 >
 >
 > Paul N. Hengen
 > National Cancer Institute
 > Frederick Cancer Research and Development Center
 > Laboratory of Mathematical Biology
 > P.O. Box B
 > Frederick, Maryland 21702-1201 USA
 > voice: (301) 846-5581
 > FAX: (301) 846-5598
 > e-mail: pnh@ncifcrf.gov
 >

 Yes There is such a dye. It is Orange G (Sigma O 1625). We use it instead
 of the bromophenolblue and xylene cyanol in Ficoll based loading dye.
 Works as a charm.

 Axel P.N. Themmen
 themmen@endov.fgg.eur.nl
From: rick@GENEMAN.WUSTL.EDU (Rick Wilson)
Newsgroups: bionet.molbio.methds-reagnts
Subject: RE: Taq errors during PCR
Date: 29 Jul 92 13:29:38 GMT
Distribution: bionet


>Does anyone know if misincorporations can be consistent?  If this was
>a problem it would be detected by sequencing the PCR product.
>
>-=] Steve [=-      AGOODRID@VAXA.WEEG.UIOWA.EDU

Pardon my incomplete response, but the answer to this question seems
to be yes.  After sequencing quite a bit of cloned PCR product, we've
observed that most Taq misincorporations seem to be T in place of C
(or was it C in place of T?, lost my notes & my memory!), or on the
complementary strand, A in place of G (with the same lost note/memory
caveat).  Anyways, consistent errors (inconsistencies??) seem to
predominate.  Hope this helps.

R.Wilson
St. Louis