Date: Thu, 09 Jan 1997 15:33:47 -0700 (MST)
From: "Terry Mulcahy 272-5792; fax 272-9107" <TMULCAHY@medusa.unm.edu>
Subject: DNA Synthesis
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Date: Mon, 28 Oct 1996 09:43:24 -0800 (PST)
From: Barbara Revere <revereb@ava.BCC.ORST.EDU>
Subject: Re: Trityl monitoring
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To: "Terry Mulcahy (505)277-5792; Fax: 277-5389" <TMULCAHY@MEDUSA.UNM.EDU>
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Hi, we use our auto analysis routinely to monitor synthesis.
We do see the drop off of trityl in the first 10-15 fractions, but then
they gradually creep up to 98-99%. I sometimes get a 100% as well
and don't have an explanation. In our old 380B we also see a drop in the
first few. I think this is the selecting out of any 'bad
sights'--non-optimum sights that are bound to occur in the start of any
synthesis. These all get capped eventually and the synthesis 'settles down'.
Why you see it drop off again I don't know. We do not make any greater
than 50mers in the 40nmol scale, as we do find that the noise vs signal
in the 40 nmol scale is very low ratio, and often we will get a zero ASWY.
Have you taken some of your low yield syntheses and gel analyzed them?
That would be a good step to finding out if your analysis is working.
We have found that in our automated sequencing facility, that oligos
with a ASWY of 94% have given great sequencing result with absolutely NO
purification.
Good luck!
Barbara Revere *revereb@bcc.orst.edu
Center for Gene Research and Biotechnology
Oregon State University, Corvallis, OR 97331
Phone: 541-737-3413 Fax: 541-737-3045
On Fri, 25 Oct 1996, Terry Mulcahy (505)277-5792; Fax: 277-5389 wrote:
> Is there anyone out there using a 394 DNA synthsizer who makes use of the
> trityl monitor? Since mine was retorfittedly installed, I have seen erratic
> step-wise yields, according to the numbers. The machine has been thoroughly
> checked out, and flow/leak tested, and seems to be performing well. However, I
> paid $5000 for the trityl monitor, in hopes of using the numbers it generates
> as a small measure of quality control. The numbers strat out at 100%, drop off
> to low numbers, from 96 to 98% and then climb back up, usually falling off
> slowly, and then climbing back up at the end. There are occasonal step-wise
> yields of 100%, and sometimes, < 85% I don't know at to make of these numbers.
> The conductivity measurements confirm this erraticism. No one has been able to
> suggest a solution. I've changed the DCM, and flushed the columns with DCM,
> even letting them sit overnight, and flushing with CH3CN. Whereas I haven't
> seen the numbers drop off to below 85% as before, I still see the numbers jump
> around a lot during every sysnthsis. Any ideas? Or is the trityl monitor a
> useless appendage? Thanks for any suggestions or thoughts. TM
>
>