Oligo stability in water at -20 C

David MacHugh dmachugh at mail.tcd.ie
Mon Aug 10 12:27:03 EST 1998


Dear Chris,

Although I never had problems with untouched frozen oligos, I had similar
problems with oligo stocks I needed for ancient DNA work.  Essentially after
a small number of freeze/thaw cycles, I found that the amplification yield
dropped dramatically with ancient DNA.  I had actually noticed previous
decreases in yield from modern DNA with well-used primers, but because they
were radio-labelled microsatellites, they were still scorable and I never
really followed it up.


 I tried the 95C followed by a snap-cool and it didn't make the slightest
bit of difference - the yield was the same as products from untreated oligos
run in parallel amplifications.

In the end I talked to the oligo suppliers (Oswel) and they indicated that
for maximum stability, all oligos should be stored in H2O aliquots (say 10
per oligo) in sterile tubes and each aliquot should not be freeze/thawed
more than 3-4 times before dumping it and moving on to the next aliquot.

I have since always stored oligos like this (even for modern samples) and so
far I have not had a reoccurrence of the original problem.  

Although I don't do it, a colleague stores his working primer aliquots in TE
in the fridge and he doesn't seem to have problems with difficult templates
either.  This approach is more economical because you don't have to dump any
"tired" aliquots.

Hope this helps.

David.

PS. If you wish I can send you the original detailed correspondence I
received from the oligo manufacturer.


____________________________________________________
   David MacHugh, Ph.D.
   Archaeological Genetics,
   Smurfit Institute of Genetics,
   Trinity College,
   Dublin 2.
   Ireland.
                          Phone: (353-1)-608-2719
                          Fax:   (353-1)-679-8558
                          Email: dmachugh at mail.tcd.ie
_____________________________________________________

----------
In article <6qn0b8$b78 at scotsman.ed.ac.uk>, chrisb at hgu.mrc.ac.uk (Chris Boyd)
wrote:


>Some years ago, there was a discussion on this ng about oligos going
>off in storage. Paul Hengen wrote it up for TIBS
>(http://www-lecb.ncifcrf.gov/~pnh/papers/TIBS/jan95.html). The upshot
>was that, yes, some oligos do go off and some can be resuscitated by a
>brief 95 C heat pulse.
>
>We are currently suffering from an oligo problem such that some (too
>many in fact) oligos stored at -20 C for six months or more are found
>not to work for PCR (sometimes even when UNUSED). On HPLC analysis we
>can clearly see degradation of the enfeebled oligos. Legend has it that
>this didn't used to happen with a previous supplier's oligos. Most
>people here follow the current supplier's recommendation and dissolve
>the oligo (supplied as a dried down pellet) in water before freezing. 
>Others use TE. Could it be that water-dissolved oligos will go off
>faster than buffer-dissolved oligos (even at -20 C with no thawing) due
>to (eg) autohydrolysis? Has anyone any data on this, or any other
>thoughts about oligo stability? I see no reason why oligos that are OK
>to start with, and stored in suitable sterile enzyme-free media,
>shouldn't last for years.
>
>Best wishes,
>-- 
>Chris Boyd                      | from, but not \ MRC Human Genetics Unit,
>Christopher.Boyd at hgu.mrc.ac.uk  | on behalf of  /  Western General Hospital,
>http://www.hgu.mrc.ac.uk/Users/Christopher.Boyd \   Edinburgh, EH4 2XU, SCOTLAND



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