In reply to the query:
-----------------------------
Does anyone use buffered 70% - 90% ethanol for the preservation of
specimens for DNA-based work and, if so, with what recipe, rationale and
results?
-----------------------------

I received several replies (not listed) from readers who thought that
buffering ethanol would be a sensible and logical procedure but who had no
actual experience to contribute. There were two replies from people who do
buffer their ethanol, as follows:

1. From: "Joe Staton" <JStaton@gwm.sc.edu>

We used to add about 0.1 ml of 1/10x TE (pH 8.0) to help with this.  I
think Tim Collins taught me this.  As I understand it, many inverts release
acids when preserved which leads to DNA destruction.  Others I know change
out the EtOH after 3 days to help with the same problem.

and 2. From: "Blair Hedges" <sbh1@psu.edu>

I usually buffer my 70-80% ethanol for specimen/DNA preservation (of
reptiles and amphibians) with TE (Tris EDTA) pH 8.0.  I buy 100X concentrate
from Sigma and bring the concentration up to 1X TE, although for large
volumes (liters) it may be a weaker TE concentration.  My reason for doing
so is not so much for pH but simply as a proven DNA buffer.  Also, I use 151
proof rum (75% ethanol) instead of laboratory grade ethanol because the
latter sometimes includes a touch of methanol which could harm DNA.  I work
in the West Indies so the rum is easy to get.

---------------------------

My conclusion is that buffering the ethanol is a sensible precaution that
may be particularly valuable for calcite-shelled animals and I intend in
future to dilute ethanol with 1 x TE rather than water.

To Blair Hedges' reasons for using rum I will mention my experience of
methanol-denatured ethanol. This is the standard shipboard fixative for the
brachiopods I receive from the IRD/ORSTOM Pacific sampling campaigns, and
brachiopods preserved in this have yielded amplifiable DNA after a decade
or so at -20. The only difficulty so far has been with some crinoids that
contain quinone pigments. These (I assume) converted the 10% methanol to
formaldehyde, the vapour of which caught me unawares when repacking the
specimens in an enclosed, hot environment..

Thank you all.

BLC

Dr B. L. Cohen
Molecular Genetics
University of Glasgow,
Pontecorvo Building,
56 Dumbarton Rd
Glasgow G11 6NU
Scotland, UK.

Phone: (+44) (0)141 330 5103 (direct line)
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                                339 8855 (switchboard)
Fax:                            330 4878
http://www.gla.ac.uk/ibls/molgen/staff/cohen-bl.html

"B.L.Cohen" <b.l.cohen@bio.gla.ac.uk>