DNA stuck in wells!

SerAln SerAln at yahoo.com
Fri Oct 25 08:14:43 EST 2002


Searching the archives of this group, I found a post from Jim Kami (200'/02/18, 
replying one of my posts) pointing to this paper:

Benore-Parsons, Marilee; Ayoub, Melissa A..  Presence of RNase A causes aberrant 
DNA band shifts.       Biotechniques, v.23, n.1, 1997.:128-131.

     Abstract:    RNase A, which is routinely added during DNA purification to 
reduce
contaminating RNA, causes shifting of DNA bands in agarose gets. DNA band sizes 
on agarose
gels increase as much as 10%-20% when RNase A is present. The low concentrations 
of RNase A
typically used to purify DNA cause shifting of select DNA bands, in contrast to 
higher
concentrations of RNase A where all bands are shifted and smeared The binding of 
RNase A to
the DNA is specific and the degree of the shift varies; not all DNA bands are 
retarded, and
the deviation is more pronounced in certain buffers. Other proteins, such as 
bovine serum
albumin or proteinase K, do not induce DNA band shift, suggesting the interaction is
specific. The binding of RNase A to DNA is reversible. The formation of 
RNase:DNA complexes
may affect experiments involving DNA:protein interactions such as gel shift, 
footprinting and
filter binding assays. Researchers performing DNA characterization from miniprep 
protocols
should be aware that RNase may cause the apparent sizes of DNA fragments to be 
altered and
obscure the presence of very small cloned fragments.

(sorry for the line wrapping)

Sergio




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