XhoI problems/Footprinting

un691cs at genius.embnet.dkfz-heidelberg.de un691cs at genius.embnet.dkfz-heidelberg.de
Wed Jun 17 09:17:02 EST 1998


On Tue, 16 Jun 1998 13:35:13 +0100, Greg  said:

>I am having the most ridiculous problems in preparing large amounts of
>small DNA fragments for DNA footprinting. I am trying to bulk up a 340
>bp fragment from one of our well used plasmids. The method is simple,
>cut SmaI, isolate by agarose gel purification a 2000bp frag (by qiagen
>qiaexII), cut XhoI, and agarose gel purify again, pulling out a 340 bp
>band. Sounds straight forward. I cannot get complete digestion with XhoI
>(~50%), which is dramatically reducing my yield. 

Re-reading your message, I would presume that your DNA is contaminated
with something in your agarose. This something is co-purified with
with the DNA with your qiaexII. Can you change to a higher grade
agarose ? can you skip the gelpurification step ?


clemens




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