Gel Shift Assay--Probe in Wells
smrodems at students.wisc.edu
Tue Aug 9 00:09:44 EST 1994
In article <325p0v$h5p at usenet.INS.CWRU.Edu>, pxb26 at po.CWRU.Edu (Peter
> What are the some common causes for probe not leaving the wells in gel shift
> What are some suggestions for avoiding this problem?
One obvious suggestion is to be sure to use a nonspecific DNA like poly
(dI-dC) or poly (dA-dT).
I have found that the choice of probe makes a lot of difference. In the
past I have tried probes that contained "extra" sequences derived from the
Bluescript plasmid. The multiple cloning site sequences seemed to cause a
lot of problems when I used crude nuclear extracts. If my desired DNA
sequences also contained Bluescript MCS sequences all the probe stuck in
the well. No amount of poly (dI-dC) could get that probe out of the well
either. However, the identical desired sequence without the Bluescript
sequences worked much better.
There have also been times when I have gotten a little bit of probe stuck
in the wells with no correlation to anything! Sometimes I think this just
happens if the moon isn't properly aligned with Venus. :-)
Steve "Some day I will get the hell out of Wisconsin" Rodems
"Then I am here for the Lee family renioun ... shur-wajo-shur"
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