High Salt RE buffers are making my gels weird
A. F. Cockburn
afc at nervm.nerdc.ufl.edu
Mon Apr 3 09:07:06 EST 1995
anderson at pharmdec.wustl.edu (Eric C. Anderson) wrote:
> here's something that i've never encountered before, just wondering if
> anyone else has or could perhaps explain why it's happening.
> when i do a plasmid digest with NcoI, EcoRI, ClaI or NotI in either the
> Clontech or Boehringer Mannheim high salt buffers (50mM Tris-Hcl, 10mM
> MgCl2, 100mM NaCl, 1mM DTT) and run them out on a gel, they run very
> strangely. the bands have a tendency to "frown" and the bands streak and
> sort of break up. when i run the exact same plasmids cut with AflII (which
<deleted to save bandwidth>
> thanks in advance,
> eric c. anderson
> anderson at pharmdec.wustl.edu
> dept. of molecular bio. and pharm. (314)362-3963 (lab)
> washington univ. school of medicine (314)862-2435 (home)
> 660 s. euclid box 8103 (314)362-7058 (FAX)
> st. louis, mo 63110
Running DNA in high salt causes it to run slower (because there are more
ions to carry the current. As the salt diffuses away, the DNA runs
faster- and this occurs at the lane edges first, causing frowning.
High salt can also cause DNA to streak, which I think is due to local
precipitation. This can also be caused by high DNA concentration or
very long DNA- you may notice that the larger bands look terrible but
smaller bands look OK.
To solve the problem, ethanol precipitate and redissolve.
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