agarose gel electrophoresis

Bernard Murray bernard at elsie.nci.nih.gov
Sat Mar 22 00:25:05 EST 1997


In article <5guhto$c3k$1 at mark.ucdavis.edu>, kcsnowden at ucdavis.edu says...

Yes, I agree with Kim.  In fact I took one of the excised bands and
stained it in ethidium bromide separately and found the whole gel
thickness was then positive so I assume it is slow penetration into the
gel (or slower than it takes to saturate the first "layer" of DNA
containing agarose.  Certainly, increasing the staining time from
eg. 10 minuted to 1 hour increases the intensity of staining (as long
as the background doesn't get too bad).
	A colleague of mine used to slice out the unstained middle
layer of preparative gels and used to get a much lower yield of
extracted DNA than me (who uses the whole layer).
		Bernard

Bernard Murray, Ph.D.
bernard at elsie.nci.nih.gov (National Cancer Institute, NIH, Bethesda MD, USA)


>I think it's probably because your gel isn't stained the whole way
>through.  If you added Ethidium bromide to your sample before loading
>(not that I think you should do this, especially if you want to
>isolate the DNA) then the band might look more homogeneous.
>Alternatively you could stain for longer or use thinner gels.
>Kim

>arump at imb-jena.de (Andreas Rump) wrote:
>>Dear Netters,
>>I have observed the following phenomenon several times: when I cut 
>>out a piece of agarose gel containing a single DNA band and place this
>>piece of gel on it's side, then I see two bands.
[SNIP]




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