HDTenEightyP/NetNews huggingface dataset:
Mon, 28 Sep 1992 10: 56:18 +0100
               Mon, 28 Sep 1992 10: 55:17 +0100
               Mon, 28 Sep 1992 13: 56:22 +0100
From: suter@VAX.MPIZ-KOELN.MPG.dbp.de
Newsgroups: bionet.molbio.methds-reagnts
Subject: pcr gems, why use them ?
Date: 28 Sep 92 12:56:22 GMT
Distribution: bionet
Content-Identifier: 12
Original-To: methods@uk.ac.daresbury
Conversion: Prohibited


Subj:	pcr hots start
--------------------------------------------------------------------------------
Daniel kim wrote:
+>With the use of Paraplast hot starts may become economically feasible for
+>everyday work.  However, I understand that one has to use at least 100 ul
+>for proper mixing of samples.  If this is the case, quite a bit of $$$$
+>will go to Taq.  Has anybody tried smaller volumes?
+I do hot starts for "minipreps" of lambda-ZAP clones.  This relieves me of
+extraneous bands.  I use paraffin wax that I got from the local Safeway, sold
+for jelly canning.  The volume I  have been using is 50 microliters total with
+about 20 microliters of pre-melted wax applied to the bottom mixture by
+pipettor.  This seems to work fine for me, but I am not doing anything that is
+particularly sensitive to reaction conditions.  With "regular" PCR (cold
+starts?) I can bring the reaction down to as little as 10 microliters and still
+have visible bands on the gel. I have never hot-started a 10 microliter PCR
+reaction, so I don't know if the lower limit can be reduced from 50
+microliters.
+
+Daniel Kim

am i missing something in this discussion ? what, in fact is a hot start ?
what i usually do is to assemble a 20 microliter PCR reaction at RT, excluding 
enzyme. 
this reaction is then heated to 95 C for 5 minutes to denature dna and then
the temp is lowered to 65 C (this all happens in the machine). i then stick
my pipet tip with 0.5 microliters thru the oil (white crude oil, sigma) and
let the cycles begin (without mixing).
is this a hot start ? what makes these pcr gems so special in comparison to
my method ? or do they contain a secret ingredient ?

cheers,

clemens
===============================================================================
Clemens Suter-Crazzolara, PhD
Max-Planck-Institut fuer Zuechtungsforschung
Abteilung Genetische Grundlagen der Zuechtungsforschung
Carl-von-Linne Weg 10
5000 Koeln 30
Tel. xx49-221-5062.221           fax. xx49-221-5062.21
suter@vax.mpiz-koeln.mpg.dbp.de
===============================================================================
       all job offers should include a self-addressed, stamped envelope
===============================================================================
Organization: Queen's University at Kingston
Date: Thu, 1 Oct 1992 11:25:57 EDT
From: <NORVALC@QUCDN.QueensU.CA>
Newsgroups: bionet.molbio.methds-reagnts
Subject: CHAOTIC PCR

   I am frustrated by what I believe is a common experience to PCRers.  I
amplify mtDNA segments from total genomic DNA.  I follow a ds amplification by
ss amplification, clean up and sequencing with Sequenase.  However, apparent
successful amplification and cleanup of the desired fragment, as evidenced
by test gels, is not a good predictor of the eventual success in sequencing
the product.  I find this extremely frustrating.
   As we all know, many variables can affect the outcome of PCR (ie. primers,
contamination, reagent conc. etc.).  Even when I perform PCR under seemingly
identical conditions, I cannot guarantee consistent success.  My questions to
the net are...
1)  What is the most critical determinant of sequencing success in PCR and
what are teh most useful predictors of success?
2)  If there are no useful predictors, is this because the steps involved in
PCR are a higher order process, and am I seeing CHAOS in action?
   I hope that the latter is not the case.  I would like to achieve consistent
results.  I'll provide a summary if this generates a discussion.  Thanks in
advance.

Barry Norval Campbell
Queen's University
Kingston, Ontario, CANADA
From: kristoff@net.bio.net (David Kristofferson)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Re: FAQ!!
Date: 1 Oct 92 22:54:13 GMT
Distribution: bionet
Organization: BIOSCI International Newsgroups for Biology

afc@gnv.ifas.ufl.edu writes:

>In article <4001@fcs280s.ncifcrf.gov>, pnh@fcsparc6.ncifcrf.gov (Paul N Hengen) writes:
>> 
>> Due to the recurrence of the chloramphenicol question,
>> I've started the construction of a FAQ list for
>> bionet.methds-reagnts and would like other people to
>> contribute to it's growth and evolution. Please feel free
>> to include what you like and any suggestions for a better
>> FAQ list. Thank You. -Paul.

>Finally, people are getting the idea that they should post summaries of
>answers when they ask a question.  Is anyone archiving these?  If so,
>an index and FTP address would be wonderful.

We will put up FAQs for FTP on net.bio.net and will have a complete
FTP and WAIS archive for all BIOSCI/bionet groups running pretty soon.
A FAQ for METHODS is a monumental project and my hat is off to Paul.

				Sincerely,

				Dave Kristofferson
				BIOSCI/bionet Manager

				kristoff@net.bio.net
From: muriana@aclcb.purdue.edu (Peter M. Muriana)
Newsgroups: bionet.molbio.methds-reagnts
Subject: Photographing Coomassie-stained Gels
Date: 29 Sep 92 15:51:08 GMT
Reply-To: muriana@aclcb.purdue.edu
Organization: Purdue University AIDS Center

Dear Netters,

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 (in regard to bracketed 
		exposure settings to see effect of lighter/darker exposure 
		since autoexposure print processing would make all of your 
		bracketed shots look the same)			


we are familiar with the Kodak self-processing system but want negatives of 
the gel as well.

Thanks in advance.   -Peter
Newsgroups: bionet.molbio.evolution,sci.bio
From: toby@carson.u.washington.edu (Toby Bradshaw)
Subject: Re: Selection for nonzero mutation rate
Organization: University of Washington, Seattle
Date: Thu, 1 Oct 1992 23:17:15 GMT

Thanks to those who replied to my inquiry on the history
of experimentation on the fitness consequences of increased/decreased
mutation rates in bacteria.  The reference I'm sure I had in mind
was provided by Barry Hall, who (along with others) noted that
fitness was increased with increased mutation rates, not decreased
by decreased mutation rates as I had (mis)remembered and posted.
The citation that follows contains references to the genetic control
of mutation rate, including reduced mutation rate in T4.  Several
people who replied to my post referred to the work in T4.  I
won't post those references; they're available in the key
paper below.

Cox, Edward C. & Gibson, Thomas C. (1974) Selection for high
	mutation rates in chemostats. Genetics 77:169-184.

	"Competition experiments in chemostats show that mutator
populations of Escherichia coli are more fit than wild type.  The
increased fitness can be explained by the appearance of new mutants
better adapted to the chemostat environment.  Fitness values vary
between chemostats and are strongly correlated (P<0.001) with
fluctuations in population density."

Thanks again to everyone who replied.

Toby Bradshaw
Department of Biochemistry and College of Forest Resources
University of Washington, Seattle
toby@u.washington.edu
From: XIAOW@SASK.USASK.CA (Wei)
Newsgroups: bionet.molbio.evolution
Subject: Re: Selection for nonzero mutation rate
Date: 1 Oct 92 18:57:00 GMT
Distribution: bionet

Bacteriophage T4 DNA polymerase mutations that affect spontaneous mutation
rates have been extensively studied by Linda Reha-Krantz and her colaborators.
Mutations deficient in 3'-5' exonuclease activity dramatically increase
spontaneous mutation and the results are in PNAS 88:2417 (1991).  I heard her
recent study on antimutator mutants of T4 DNA polymerase.  People who are
interested in this issue may have to contact Linda for detail.
Wei Xiao
Department of Microbiology
University of Saskatchewan
Saskatoon, SK, Canada S7N 0W0
From: bickle@urz.unibas.ch
Newsgroups: bionet.molbio.evolution
Subject: Re: Selection for nonzero mutation rate
Date: 1 Oct 92 07:47:36 GMT
Followup-To: bionet.molbio.evolution
Organization: University of Basel, Switzerland

In article <4003@fcs280s.ncifcrf.gov>, toms@fcs260c2.ncifcrf.gov (Tom Schneider) writes:
> Bacteriophage T4 DNA polymerase apparently has mutations which decrease the
> rate of mutation formation.  So mutation rate is under genetic control.  Sorry,
> I don't know a direct referece.
> 

The T4 DNA polymerase has mutations which work both ways: antimutators which
decrease mutation rates have either increased proof-reading 3'-5'
exonuclease activity or reduced affinity for the wrong precursors.
Mutators with *increased* mutation rates have, as I recall, a decreased
proof-reading activity.

See chapter 4 in DNA replication and mutagenesis (R.E. Moses and
W.C. Summers eds.) ASM 1988 ISBN 1-55581-003-9 for a reasonably recent 
review.

Tom Bickle
Dept of Microbiology
Biozentrum, Basel U.
Basel Switzerland  
Xref: sparky bionet.molbio.evolution:350 sci.bio:3401
From: toby@milton.u.washington.edu (Toby Bradshaw)
Newsgroups: bionet.molbio.evolution,sci.bio
Subject: Selection for nonzero mutation rate
Date: 28 Sep 92 17:38:32 GMT
Article-I.D.: u.1992Sep28.173832.20045
Reply-To: toby@u.washington.edu
Organization: University of Washington, Seattle

I vaguely remember some experiments showing that
bacteria with abnormally low rates of mutation
are less fit than those with "normal" levels.  
Does anyone have references to this line of investigation?
I'll be happy to post a summary of responses.

Toby Bradshaw
Department of Biochemistry and College of Forest Resources
University of Washington, Seattle
toby@u.washington.edu
Xref: sparky bionet.molbio.evolution:351 sci.bio:3426
From: toms@fcs260c2.ncifcrf.gov (Tom Schneider)
Newsgroups: bionet.molbio.evolution,sci.bio
Subject: Re: Selection for nonzero mutation rate
Date: 30 Sep 92 18:01:16 GMT
Followup-To: bionet.molbio.evolution
Organization: Frederick Cancer Research and Development Center

Bacteriophage T4 DNA polymerase apparently has mutations which decrease the
rate of mutation formation.  So mutation rate is under genetic control.  Sorry,
I don't know a direct referece.

  Tom Schneider
  National Cancer Institute
  Laboratory of Mathematical Biology
  Frederick, Maryland  21702-1201
  toms@ncifcrf.gov
Xref: sparky bionet.molbio.embldatabank:44 bionet.software:1843
Newsgroups: bionet.molbio.embldatabank,bionet.software
Subject: EMBL-Search v2.1 available
From: Fuchs@EMBL-Heidelberg.DE (Rainer Fuchs)
Date: 28 Sep 92 10:41:35 +0100
Distribution: world
Organization: EMBL Data Library

Version 2.1 of EMBL-Search is now available.

EMBL-Search is a free database query and retrieval program for the Apple
Macintosh for use with the EMBL Sequence Databases CD-ROM. Version 2.1
allows text queries of the PROSITE protein pattern database, in addition to
the EMBL nucleotide and SWISS-PROT protein sequence databases.
Cross-references between EMBL, SWISS-PROT, PROSITE (data) and PROSITE (doc)
can be used to navigate easily between these databases.

EMBL-Search requires the December 1991 EMBL CD-ROM or later. For queries of
PROSITE, the new September 1992 CD-ROM (Disk 1) is needed.

EMBL-Search 2.1 will be provided in binhexed form on the next EMBL CD-ROM.
It can also be obtained by electronic mail from NetServ@EMBL-Heidelberg.DE
(get mac_software:embl-search.hqx) or by anonymous ftp from
ftp.embl-heidelberg.de (/pub/software/mac/embl-search.hqx). The complete
source code is available as well (embl-search_src.hqx).

Rainer Fuchs
EMBL Data Library
Fuchs@EMBL-Heidelberg.DE
From: wixer!will@cs.utexas.edu (Bill Staniewicz)
Newsgroups: bionet.molbio.embldatabank
Subject: request info about the list
Date: 30 Sep 92 17:35:07 GMT
Distribution: bionet
Original-To: embl-db@uk.ac.daresbury
X-Mailer: ELM [version 2.3 PL11]

Hi,

As a new subscriber, I would be interested
information about the list. Could someone post a brief
summary or email me directly?

Thank you.

-- 
Bill
will@wixer.cactus.org
From: Fuchs@EMBL-Heidelberg.DE (Rainer Fuchs)
Newsgroups: bionet.molbio.embldatabank
Subject: EMBL anonymous ftp server - please use correct address
Date: 5 Oct 92 08:38:51 GMT
Followup-To: bionet.molbio.embldatabank
Organization: EMBL Data Library

There seems to exist some confusion concerning the correct address of the
EMBL anonymous FTP server. Some users complained that they could not see
any files and could not change directories.

The simple reason was that an incorrect Internet address was used. The EMBL
anonymous FTP server is located at ftp.embl-heidelberg.de [192.54.41.33],
and
NOT embl-heidelberg.de. The latter is a completely different machine.

Secondly, our Internet connection is not yet optimal despite our attempts
to
improve the situation. As Jaime Prilusky has pointed out in a previous
posting
to this newsgroup, the Israeli EMBnet node at sunbcd.weizmann.ac.il mirrors
the EMBL archives and might provide better connections for some users.

Rainer Fuchs
EMBL Data Library
Fuchs@EMBL-Heidelberg.DE
Newsgroups: bionet.molbio.embldatabank
Subject: Slow ftp.embl-heidelberg.de
From: Jaime Prilusky <lsprilus@weizmann.weizmann.ac.il>
Date: Sat, 26 Sep 1992 18:04:06 GMT
Organization: INN - Weizmann Institute of Science - Israel
X-Useragent: Nuntius v1.1

In article <Fuchs-250992081601@rainer-mac.embl-heidelberg.de> Rainer
Fuchs,
 Fuchs@EMBL-Heidelberg.DE writes:
>The simple reason is that the German networks and their connection to the
>international Internet are not very effective, especially for generators
of
>huge amounts of traffic such as EMBL.
>
>We are very actively investigating different approaches to improve this
>unsatisfactory situation.

Please remember that our anonymous ftp server holds a COMPLETE COPY of
both
SOFTWARE AND DATA of the  ftp.embl-heidelberg.de server, and our
connection
access time is really good.

Just anonymous ftp to         sunbcd.weizmann.ac.il
                              132.76.64.79

You can also access our data and software into our INN Gopher, to the
same address
and port 70.

 Dr Jaime Prilusky
 Israel National Node   INN          ! LSPRILUS@WEIZMANN.WEIZMANN.AC.IL
 Weizmann Institute of Science       ! fax: 972-8-344113
 76100 Rehovot - Israel              ! tel: 972-8-342470 / 2979
Newsgroups: bionet.molbio.embldatabank
Subject: EMBL CD-ROM goes double
From: stoehr@embl-heidelberg.de
Date: 25 Sep 92 19:52:42 +0100
Organization: EMBL, European Molecular Biology Laboratory

The September 1992 issue of the EMBL CD-ROM will consist of two disks for the
first time. We have arranged directories and files so that typical things can
still be comfortably performed if you just have one CD-ROM drive.
However, those people who use the EMBL CD-ROM on file servers may need this
advance warning to rearrange their activity.

Existing or prospective subscribers for the 1992/3 susbcription period need do
nothing extra - the prices are the same - you will get both disks together. We
expect to distribute this issue in 7-10 days.

On volume 1, we include:
- EMBL and Swiss-Prot in their flat-file distribution format
- the MS-DOS and Mac query retrieval software and indices (which 
  operate on the above flat-files)
- related databases, particularly those with cross-references to/from the
  sequence databases (eg EPD, ECD, Prosite, Enzyme, REBASE)

On volume 2, we include:
- EMBL and Swiss-Prot in a 'FASTA' format, for use with Bill Pearson's
  FASTA for Mac or PC.
- EMBLSCAN PC program and indices for fast lookup for very similar sequences
  in the EMBL nucleotide database.
- a whole host of other molecular biology databases
  (eg TFD, FlyBase, Kabat, Berlin RNA database, RLDB, Seqanalref, tRNA, BLOCKS)

Regards,
Peter Stoehr
EMBL Data Library
Newsgroups: bionet.molbio.embldatabank
Subject: MacPattern (Was: ftp at ftp.embl-heidelberg.de
From: Fuchs@EMBL-Heidelberg.DE (Rainer Fuchs)
Date: 28 Sep 92 10:28:02 +0100
Followup-To: bionet.molbio.embldatabank
Distribution: world
Organization: EMBL Data Library

In article <1992Sep24.005109.21258@nmr-z.mgh.harvard.edu>,
cherry@OPAL.MGH.HARVARD.EDU wrote:

> The ftp.embl-heidelberg.de ftp site is on a very slow network link to
> the outside world. You can retrieve MacPattern from any of the hosts
> listed below. This is what archie says is available on macpattern.

The list that followed in this message showed one of the weaknesses of
Archie:
you can't find information about the authorative site for a program. All
sites listed had outdated versions, sometimes about 2 years old. I don't
want to blame Archie; it can't store more information than is provided. The
Internet IAFA working group is currently trying to establish standards for
describing ftp archives and their contents which will include "authority"
information that could then be picked up by Archie in the future. 

> The EMBL mail server is still one of the best ways to retrieve files
> from Heidelberg.

Nothing to add to this. We keep trying to improve our Internet links.

Rainer Fuchs
EMBL Data Library
Fuchs@EMBL-Heidelberg.DE
Organization: University of Illinois at Chicago
Date: Friday, 25 Sep 1992 22:02:34 CDT
From: Yakof <U50701@uicvm.uic.edu>
Newsgroups: bionet.molbio.hiv
Subject: Re: General information inquiring about HIV
Distribution: bionet

1 The virus will be transfered  only with blood in this case, so one would have
 to cut self with a shaver that just cut HIV patient.


2 There are experiments that can show that it [virus] can survive in the air fo
r up to 24 hrs.


3 Soap and alcohol cannot garantee that.  Bleach is the best agent to use...




4 Gneral means of contracting HIV is through body fluids: blood, sperm, saliva,
 serum etc.  Though, saliva has a very low HIV/mL and is a very unlikely way of
 contracting HIV.  But you still want to be carefull, the donts then are:
  a. unprotected sex (either homo or heterosexual)
  b. sharing of IV needles
  c. getting untested blood (thru IV)
  d. oral sex without protection
  e. placenta is not a barrier for HIV


any questions? you can write me or post a note. take care.
From: karabinos@exmed1.dnet.gwdg.de ("EXMED1::KARABINOS")
Newsgroups: bionet.molbio.hiv
Subject: sub
Date: 28 Sep 92 08:49:03 GMT
Distribution: bionet
Original-To: hiv-biol@uk.ac.daresbury

sub bionet.molbio.hiv
Newsgroups: bionet.molbio.hiv
From: biz@columbia.Princeton.EDU (Dave Bisignano)
Subject: Just a Question
Originator: news@nimaster
Reply-To: biz@soil.Princeton.EDU
Organization: Princeton University
Date: Tue, 6 Oct 1992 18:54:46 GMT


Do condoms (latex) have pores in them,
 if so how large are they as opposed to the hiv virus? 
 I have recently heard that there are holes
 and they are 7 times larger than the virus.
Organization: University of Illinois at Chicago
Date: Friday, 25 Sep 1992 21:57:55 CDT
From: Luiz  <U50701@uicvm.uic.edu>
Newsgroups: bionet.molbio.hiv
Subject: Re: General information inquiring about HIV
Distribution: bionet


The virus cannot be 100% killed by regular soap.
Clorine does a better job (100% effective)
From: GenBank-Updates@genbank.bio.net
Newsgroups: bionet.molbio.genbank.updates
Subject: Simian immunodeficiency virus (M87004) glycoprotein 120 (env) gene.
Date: 29 Sep 92 10:09:22 GMT
Distribution: bionet
Checksum: 65041     2

LOCUS       SIVM7004AA    258 bp ds-DNA             VRL       29-SEP-1992
DEFINITION  Simian immunodeficiency virus (M87004) glycoprotein 120 (env) gene.
ACCESSION   M96197
KEYWORDS    envelope protein; glycoprotein 120.
SOURCE      Simian immunodeficiency virus (individual_isolate Mne) DNA.
  ORGANISM  Simian immunodeficiency virus
            Viridae; ss-RNA enveloped viruses; Positive strand RNA virus;
            Retroviridae; Lentivirinae.
REFERENCE   1  (bases 1 to 258)
  AUTHORS   Overbaugh,J. and Rudensey,L.M.
  TITLE     Alterations in potential sites for glycosylation predominate during
            evolution of the simian immunodeficiency virus envelope gene in
            macaques
  JOURNAL   J. Virol. 66, 5937-5948 (1992)
  STANDARD  simple staff_entry
BASE COUNT       60 a     59 c     45 g     94 t
ORIGIN
        1 taccacctct gcactcatgg tgatattagt ttggtttcca tcaatccaat ctatgtttgc
       61 tatgagactg gtcactgtgg aattacacgt gaggtctccc tctcttggag gcaaataaac
      121 atttctgcct actttatgcc aagtgttgat tatttgtcta atatgacatg gcacgtaatt
      181 ccttttatgc cgttcctgtg gcttctgggt agttccattc agatccttat cttctaccca
      241 atttagaaac caattcat
//
From: GenBank-Updates@genbank.bio.net
Newsgroups: bionet.molbio.genbank.updates
Subject: Simian immunodeficiency virus (F87038) glycoprotein 120 (env) gene.
Date: 29 Sep 92 10:08:53 GMT
Distribution: bionet
Checksum: 13434     2

LOCUS       SIVF87038C    258 bp ds-DNA             VRL       29-SEP-1992
DEFINITION  Simian immunodeficiency virus (F87038) glycoprotein 120 (env) gene.
ACCESSION   M96178
KEYWORDS    envelope protein; glycoprotein 120.
SOURCE      Simian immunodeficiency virus (individual_isolate Mne) DNA.
  ORGANISM  Simian immunodeficiency virus
            Viridae; ss-RNA enveloped viruses; Positive strand RNA virus;
            Retroviridae; Lentivirinae.
REFERENCE   1  (bases 1 to 258)
  AUTHORS   Overbaugh,J. and Rudensey,L.M.
  TITLE     Alterations in potential sites for glycosylation predominate during
            evolution of the simian immunodeficiency virus envelope gene in
            macaques
  JOURNAL   J. Virol. 66, 5937-5948 (1992)
  STANDARD  simple staff_entry
BASE COUNT       60 a     58 c     46 g     94 t
ORIGIN
        1 taccacctct gcactcatgg tgatattagt ttggtttcca tcaatccaat ctatgtttgc
       61 tatgagactg gtcactgtgg aattacacgt gaggtctccc tctcttggag gcaaataaac
      121 atttctgcct actttatgcc aagtgttgat tatttgtcta atatgacatg gcacgtaatt
      181 ccttttatgc cgttcctgtg gcctctgggt agttccagtc agattcttat cttctaccca
      241 atttagaaac caatttat
//
From: GenBank-Updates@genbank.bio.net
Newsgroups: bionet.molbio.genbank.updates
Subject: Simian immunodeficiency virus (F87189) glycoprotein 120 (env) gene.
Date: 29 Sep 92 10:09:00 GMT
Distribution: bionet
Checksum: 45097     2

LOCUS       SIVF87189D    203 bp ds-DNA             VRL       29-SEP-1992
DEFINITION  Simian immunodeficiency virus (F87189) glycoprotein 120 (env) gene.
ACCESSION   M96184
KEYWORDS    envelope protein; glycoprotein 120.
SOURCE      Simian immunodeficiency virus (individual_isolate Mne) DNA.
  ORGANISM  Simian immunodeficiency virus
            Viridae; ss-RNA enveloped viruses; Positive strand RNA virus;
            Retroviridae; Lentivirinae.
REFERENCE   1  (bases 1 to 203)
  AUTHORS   Overbaugh,J. and Rudensey,L.M.
  TITLE     Alterations in potential sites for glycosylation predominate during
            evolution of the simian immunodeficiency virus envelope gene in
            macaques
  JOURNAL   J. Virol. 66, 5937-5948 (1992)
  STANDARD  simple staff_entry
BASE COUNT       42 a     37 c     37 g     87 t
ORIGIN
        1 tttatcattg gctcttgttc caagcctgtg caattatcat gatttataca agaactattc
       61 tcattgacca cttctattgc ctctgttgtt gttggtgttt ctgttggtgt tgttgttgtt
      121 gatgattttg tcaatcccca tctatctgtc tcacttttgt tgcatctcat agaaatgcat
      181 aatggggtta attttacaca agg
//