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RNA denature gel speed?


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#1 Tintin

Tintin

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Posted 10 August 2009 - 12:16 PM

I ran an 8cm RNA denature gel in 6.5V/cm (is that too high? my gel shrink a little again, about at the 2mm end of the gel). I found that the bromophenol blue run 3/4 within one hour (6cm/hour). Is that too fast? I remember that it should take about 3-4 hour for the dye to run 6cm. Is my gel wrong or my memory? Any one can tell me the right speed of the migration?

My gel recipe: 50mM boric acid, 1mM sodium citrate, 5mM NaOH, pH7.5, 0.3M formaldehyde.

Loading buffer : 50mM boric acid, 1mM sodium citrate, 5mM NaOH, pH7.5, 50% formalmide, 1.85M formaldehyde, 5% glycerol, bromophenol blue, 0.1M EDTA pH8, 50ng/ul EtBr

Running buffer:50mM boric acid, 1mM sodium citrate, 5mM NaOH, pH7.5

I just tried with some DNA ladder this time rather than my real RNA sample. I denatured the DNA as the RNA denature procedure (mix with " 50mM boric acid, 1mM sodium citrate, 5mM NaOH, pH7.5, 50% formalmide, 1.85M formaldehyde", 65 degree 15min). I'll upload the pic later.




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