XhoI problems/Footprinting

Ashok Aiyar aiyar at ebv.oncology.wisc.edu
Tue Jun 16 08:14:06 EST 1998


On Tue, 16 Jun 1998 13:35:13 +0100,
Greg (g.dean at uea.ac.uk) wrote:

>If you have any ideas about my XhoI problems or suggestions for
>purifying large amounts of small fragments of DNA from plasmids for
>footprinting, I would be interested in hearing them.

I don't have any suggestions vis-a-vis XhoI, but as an alternative
method of making your 340 bp fragment why don't you simply amplify
it using PCR from the plasmid.  Oligonucleotide primers are priced
low enough that it is likely that synthesizing your fragment this
way will be cheaper than cutting it out of a plasmid.  Finally to
label a specific end for PCR, all you need to do is to phosphorylate
one of the primers with gamma-32P-ATP prior to PCR.  I have used
PCR to make uniformly labeled fragments and end-labeled fragments
several times with success.

Later,
Ashok
-- 
Ashok Aiyar, Ph.D.
McArdle Laboratory for Cancer Research
aiyar at ebv.oncology.wisc.edu



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