Fuzzy Sequencing Gel Bands

Jim Owens jow at helix.nih.gov
Wed Feb 16 09:26:41 EST 1994


In article <01H8VS2QXCJ68WZ7PJ at MSSCC.MED.UTAH.EDU> ,
schoenfeld at MSSCC.MED.UTAH.EDU writes:
>Can anyone tell me why I'm getting fuzzy bands on sequencing gels?  For
>each sample, I usually do 2.5 hr and 6 hr runs side by side.  The 2.5 hr
>run looks fine, but the 6 hr run is fuzzy.  Since they're on the same
gel,
>I don't think the fuzziness is due to something like gel temperature or
>buffer depletion. Any ideas?

Not really. But I have had gels in which some areas of the autoradiogram
have fuzzy bands and others do not.  I have attributed this to "air
bubbles" between the dried gel and the film.  How this could happen I
haven't the faintest idea.  

Another possibility is slight stretching during the transfer from glass
plate to blotting paper.  This would displace the bands on one side of
the gel with respect to the other side.  

This last could also happen if the gel runs faster on one face than on
the other.  I've seen this on thicker (>=1mm), non-sequencing gels, both
polyacrylamide and agarose.

Good luck,

Jim Owens



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