ABI 310 DNA sequencer and Pop6

Phillip San Miguel pmiguel at purdue.edu
Mon Mar 13 10:04:58 EST 2000


Mick Guerini wrote:

> [...]    Secondly, we have encountered in the last 60 days a problem
> relating
> to the Pop6 chemistry.  Two lots in particular have problems and we have
> determined this by trial and error and by finding a lot that worked
> great for 105 runs (one whole bottle of polymer).  Let me explain the
> problem briefly and see if anyone out there has any suggestions or
> comments.  The problem is intermittent, 4 or 5 runs may work well then a
> run has good peaks (crisp, tight, tall) for the first 200 some odd bases
> and then the sequence loses resolution and looks like wide waves. This
> problem is intermittent in the first 10-12 runs on a new capillary and
> then the capillary goes bad.  Does anyone have this problem with Pop6
> currently (or previously)?
> [...]

    I don't have a 310, but I have this problem intermittantly in some of
the caps in my 3700. I actually didn't notice it using POP6, I use all POP5
now. Usually the resolution is bad for the entire read (and the first peak
is delayed by as much as 10 minutes with respect to the good reads.) But
I've seen at least one case of a read that started out normal and then lost
resolution.
    PE isn't much help on this one. They suggest it is the result of a
bubble migrating through the capillary. Whatever that means.
    It does seem to be template and cap specific. I see it happening more on
certain batches of templates. I'm going to try not growing our bugs as long,
see if that helps. Also, I've been loading 1/2 as much sample on the machine
(1/2 the concentration) and I think that might help.
   Another possibility: there really are bubbles in the caps sometimes.
Before I switched to the new version of data collection software for the
3700 I had the cap refill times set high (30 minutes) and didn't see this
problem (or never noticed it.) Maybe the quicker refill times I'm using now
are causing these bubbles.

>    An additional question:  Has new chemistry been developed for the
> 3700 (96 capillary machine) and does it work in the 310?
> [...]

POP6 and POP5 are the polymer chemistries available on the 3700 at the
moment. Anyway, I've seen this problem with POP5--so it is not POP6
dependant.

Phillip San Miguel
Purdue University Genomics Core Facility


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