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Electrophoretic Mobility Shift Assay Forum EMSA Forum. Discuss the Electrophoretic Mobility Shift Assay protocols or technique and the assessment of DNA and RNA binding proteins here. Also post questions and troubleshooting about the shift assay.


RNA EMSA percentage of gel

RNA EMSA percentage of gel - Electrophoretic Mobility Shift Assay Forum

RNA EMSA percentage of gel - EMSA Forum. Discuss the Electrophoretic Mobility Shift Assay protocols or technique and the assessment of DNA and RNA binding proteins here. Also post questions and troubleshooting about the shift assay.


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Old 01-12-2009, 10:17 PM
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Default Re: RNA EMSA percentage of gel

Hi Kevin_a,

I am also doing EMSA with RNA probe. I encountered the same problem which my shifted band seems stuck in the well. Since you mentioned that is also considered as binding, I am kind of wondering how can I make the stuck band move in the gel, are there any other way rather than lowing the percentage fo the gel? I will really appreciate your thought on this.

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Originally Posted by kevin_a View Post
Typical DNA EMSA use a 5 or 6% native gel with ~20bp double stranded probe.
Your's is half the mass, so doubling will help, but you'll probably still see diffusion admin pointed out.

You can't just increase the percentage of gel because your proteinrobe complexes are going be very high up on the gel - possibly even look stuck in the well. If you are looking for binding vs no binding it may not matter as much, but if you need to resolve multiple bands you will probably want to tweek with the gradients.
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Old 01-16-2009, 06:04 PM
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Default Re: RNA EMSA percentage of gel

Hi Luo,
have you tried a gradient gel?
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  #13  
Old 12-18-2009, 09:16 AM
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Default Re: RNA EMSA percentage of gel

Help Help, me with RNA EMSA.
the free RNA probe(size 1:200 nt,size 2:2000nt,size 3:1800 nt) i prepared when i run in 4% native PAGE is stucked in the bottom of the loading well. i dunot know why there is no movement even i run the gel for 3.5 hrs/120V. please help me out from this problem. i would be really thankful to get any suggestion ASP.
Thank you.
Bilon,Japan
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