ABI 310 DNA sequencer and Pop6

Gys de Jongh GysdeJongh at compuserve.com
Mon Mar 13 18:39:22 EST 2000

"Mick Guerini" <mguerini at gemini.oscs.montana.edu> wrote in message
news:8aie9s$qh1$1 at mercury.hgmp.mrc.ac.uk...

[ABI-310 performance and POP-6 problems]

we also noticed that the ABI-310 funtions better with the short capillary. The
short capillary produces the same number of bases at the same resolution in much
less time. The long capillary adds a few bases at low resolution. ( In "our
hands" of course )  If the sequence is longer than 500 bases we sequence "from
the other side". The problem with the POP-6 sounds familiar. We had exactly the
problem you describe : loss of resolution after 100-200 bases and damaging of
the capillary. This was at the end of 1998 / beginning 1999. Perkin elmer was
aware of the problem and the lot numbers of the supected POP-6 but forgot to
notify us. We too replaced the pump block , the syringe , the water etc. We
discovered what the problem was after we borrowed a batch of POP-6 from another
departement . This lot was expired for about a year but worked fine for the next
500 injections. Perkin elmer did not seem to know why this batch of POP-6 was
less optimal. The [EDTA] was suspected. I am almost sure that it was the POP-6
because first i replaced every thing except the POP-6 : the problem stayed. Than
i changed only the POP-6 and the problem was solved. The previous discussion
also points to the POP-6 :

Subject: late reply for ABI seq. problem
From: "Marcel Schijf" <M.A.Schijf at pharm.uu.nl>
Date: 1999/02/10
Newsgroups: bionet.molbio.methds-reagnts
Hit htere i had the same problem witm my sequencing reactions, whatever the
reaction i couldn't get any reads.

when i phoned PE they came by and found out that the problem lay with a bad
POP-6 polymere batch..they changed the cappillary (it was affected by the
bad lot) and changed the polymere..and voila all my reads were just

but PE didn't gava out a message thath twol lots off POP6 were bad ..i know
the last 3 digits from the bad polymere i had ..and it ended in 012...maybe
you want to check your POP6

Subject: Re: ABI 310 problems
From: "Hans van Zyl" <hvanzyl at idpi1.agric.za>
Date: 1999/01/15
Newsgroups: bionet.molbio.methds-reagnts

T.J. Young <BMBTJY at leeds.ac.uk> wrote in article
<7758el$8fk_001 at leeds.ac.uk>...

> just recently the data I was getting was rubbish, the peaks would start off
> ok but then gradually widen after about 80bp turning into complete garbage
> by 150bp :(  I've tried the obvious things:  Flushing the capillary through
> with plenty of sterile water, using fresh buffer and polymer but they made
 >no difference.

I assume you use POP polymer chemistry? We do 310 microsatellite analysis
and if we see loss of resolution that increases during the run it usually
is an indication of a shot capillary.
Try use a new capillary (part number 402839) and use the buffer with added
EDTA  (part number 402824) as this greatly enhances capillary life...

Apart from apparent manufacturing problems with the POP-6 we think the ABI is a
very good machine. From day one on we are sequencing DNA well within its spec's
, even with diluted ready reaction mix. We found the machine needs very little
hands-on time , has a very good manual , is so user friendly that guests and
students can learn to use it in a few hours.



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