Oligo stability in water at -20 C

Michael Benedik benedik at uh.edu
Tue Aug 11 11:49:05 EST 1998


In article <199808111530.JAA74240 at nestor.NMSU.Edu>
hroychow at NMSU.EDU (Hiranya Roychowdhury) writes:

> At 01:31 AM 8/11/98 GMT, Michael T. MacDonell wrote:
> >Dear Hiranya:
> >
> >Maybe.
> >I just have not observed it to be significant, and I have been
> >observing for about 15 years.  Anyway, I wonder how much spontaneous
> >degradation is actually attributable to nucleases.  How are can you be
> >sure you are not seeing nuclease degradation and believing that it to
> >be autocatalysis?
> >
> >Best Regards,
> >Mike MacDonell, Ph.D.
> >Chief Scientific Officer
> >Ransom Hill Bioscience
> >
> 
> Dear Mike,
>         While nuclease may not be totally discounted in any situation,
> autocatalysis of nucleic acid is a fact. The reason I'm sure about this is
> that I had performed a simple expt a couple of years back. I made up two
> tubes of a working conc. of a DNA size markers: one in plain water
> (nuclease-free) and the other in TE. they were both stored at 4 C in the
> walk-in cooler. I checked the markers periodically. After about six weeks,
> the markers stored in plain water started showing diffused pattern, and, by
> about another four weeks, only a smear on the gel used to be visible. The
> markers stored in TE continued to show sharp bands till the tube was empty. 
>         The reason for this demonstration was a grad student's show of
> disbelief at the suggestion of NA autocatalysis. She had made up aliquotes
> of the DNA markers in plain water (about 40 of them), and they were of no
> use after some time. The degradation is minimal at -70 C. At -20 C, unless
> undergoing repeated thawings, it proceeds but very slowly. Degradation due
> to nucleases at 4 C, or even at -20 C, does not take very long.
> with regards,
> Hiranya
> 
> 
> Dr. Hiranya Sankar Roychowdhury
> Plant Genetic Engineering Lab.
> New Mexico State University
> Las Cruces, NM 88003
> Ph. (505) 646-5785
> hroychow at nmsu.edu
> 


Hiranya

Whilst I am not an expert on autocatalysis, the expt you describe above
really doesn;t distinguish between nuclease contamination, bacterial
contamination, or autocatalysis. The TE would buffer the soln, but the
edta in TE would also protect from nucleases, whether from bacterial or
other sources. 

If your model is correct, Tris alone (without EDTA) should prevent
autolysis but have no effect on nuclease. EDTA alone should inhibit
nucleases from most sources (although I don't know what "buffering"
effect it may have).


Michael Benedik
Department of Biology and Biochemistry
University of Houston
Houston, TX 77204-5934
Tel: 713-743-8377
FAX: 713-743-8351
benedik at uh.edu



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