发信人: moso (野鬼), 信区: Biology
标 题: Re: help please, have trouble with Gelshift
发信站: Unknown Space - 未名空间 (Mon Aug 30 13:28:46 2004), 转信

It seems to me that the eletrophoresis failed to seperate the free probe from
DNA-protein complex. 400 bp probe is a lot big. I quite agree with the
suggestion of adjusting your gel concentration or Acr/Bis ratio. I think it
might be easier to use the probe only to optimize the gel first, at least to
let the free probe run to the bottom. You don't need to label the DNA. Instead
you can use EB to stain the DNA. To my personal opinion, I think 5% gel is not
the best choice. Gernerally I believe 3% even lower would be better.

Good luck !

【 在 rabbitmm (永远的红茶兔子女王) 的大作中提到: 】
Hi there! I meet trouble again. ( I really think about quit and marry
someone, since my experiment never work)

I am doing gel shift assay. I run 5% acrymide gel for about 3 hours, 200v. 0.5
TBE buffer.
but my probe seems always haning in the middle, never go down to the bottom.
so it mess up with my bands. :(

My probe is really big, around 400bp. I know its not standard, but my Shijie
use it and also get a clean picture of the shift. But I do the same experiment
3 times, always get the same problem.

really confused.......



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※ 修改:·rabbitmm 於 Aug 29 20:25:10 修改本文·[FROM: 160.94.]


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※ 来源:．Unknown Space - 未名空间 mitbbs.com．[FROM: 165.230.]