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RNA gel

RNA gel - RNA Techniques Forum

RNA gel - Post and discuss RNA methods and RNA science related topics. RNA extraction, cDNA synthesis, RNA EMSA, and other RNA protocols.


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  #1  
Old 08-29-2007, 09:42 AM
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Question RNA gel



Hi guys,

i m continuousely facing a problem while runnig RNA gel. Problem is that the bands run in opposite direction. I mean when i run the gel both dyes (xylene and bromophenol blue) run along the gel but when i view the gel under UV the bands are in opposite direction (almost at the end of -ve termini and are about to go into buffer). If i run it for longer time they goes out of the gel into the buffer. ( i checked my termini they are always at right position)

I repeated the procedure for several times with fershly isolated RNA. Used DEPC H2O wherever required. used 1.2% agarose and for some times 1.5%

plz tell me is it possible that these orange golden bands in the opposite direction are either RNA or it is only EtBr that is shining.

Another problem that is facing by a friend of mine is :her RNA bands are not resolved into 28s and 18s. Moreover she is using DNA ladder as markers and marker bands are also not resolving (she has no option other than using DNA marker).
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Old 08-30-2007, 01:14 AM
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Default Re: RNA gel

Sometimes if you load too much RNA it won't travel out of the well, check your concentrations.

Check the pH of your DEPC-treated water, low pH will increase RNA degradation, and DEPC treatment can generate carbonate products that can lower resistivity and hinder electrophoresis. Also check your TBE or TAE buffers.

As far as RNA ladders go I've heard of Fermentas' RiboRuler Rna ladder, but I've never used it.

As last resort you can try PAGE
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Old 08-30-2007, 03:47 AM
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Default Re: RNA gel

thanks for replying

Its not like that RNA does not comes out of the well it does but in oppsosite direction. If i run the gel deliberately in opposite direction dyes go into the opposite direction while rna goes into the right direction its a very strange thing bcoz both the dye and rna shud run in the same direction but it does not
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  #4  
Old 08-30-2007, 02:03 PM
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Default Re: RNA gel

Quote:
Originally Posted by Aaina View Post
thanks for replying

Its not like that RNA does not comes out of the well it does but in oppsosite direction. If i run the gel deliberately in opposite direction dyes go into the opposite direction while rna goes into the right direction its a very strange thing bcoz both the dye and rna shud run in the same direction but it does not
Some Dyes are dissolved in acidic solutions, or non-treated water, where bromophenol blue would be slightly acidic, and carry a positive charge, therefore migrating to the negative terminal, the opposite of negative charged RNA.

Last edited by danfive; 08-30-2007 at 02:11 PM.
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Old 09-08-2007, 04:19 PM
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Default Re: RNA gel

Good Morning Every One
Please Some One Will Explain Me.

How Will Show My Picture Gel Here
This Foro???

Thank You

Nice Weekend

Margarita
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  #6  
Old 09-08-2007, 04:21 PM
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Default Re: RNA gel

My Picture Is No In Url
Is In Jpg.

Thank You
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Old 01-13-2008, 05:35 AM
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Default Re: RNA gel

Relax. No, that's not RNA. It's free EtBr. You must have added EtBr in your loading buffer, did you? EtBr runs to the negative while your RNA to the positive.
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