Since my posting about SAPS and NCBE protocols for extracting and manipulating
DNA I have had a couple of queries relating to staining of DNA on gels,
particularly from Barbara Moffat (whose transmission didn't contain an Email
address) - if you're out there Barbara....
WE use either
A) Methylene blue (0.1% in 0.5M Na acetate) and stain gels
(ideally) overnight and then destain with one rinse of tapwater and then
leave in fresh tapwater for about an hour - gels will keep for months if kept
in the resulting dilute stain, particularly if refrigerated. OR
B) Azure A (0.08% in 40% IMS) - the stain is poured onto the surface of the
gel and left for 4 min. Stain is then poured off and reused. The surface of
the gel is then rinsed with 70% ethanol for a few seconds and this is then
discarded. The gel is then rinsed three or four times with tap water. The
stain migrates into the gel and bands will be visible after 10min, but best
results are obtained by leaving the gel in a sealed container to prevent
drying out overnight.
When I started this DNA work I used Meth blue, but in our workshops for
teachers I now use the Azure A because it's faster.
Sensitivity is about 20% that of ethidium bromide but with the amounts of DNA
we use (0.5 to 1.0 microgram for each digest) this is not a problem.
Hope this helps.
Barry Meatyard, SAPS Programme, Inst of Ed, University of Warwick.