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Problems with Electrophoresis of RNA

RNA electrophoresis plant RNA band resolution

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#1 Flopilus

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Posted 26 September 2012 - 08:58 AM

I run a 1% agarose gel for my samples of plant RNA in 2 occasions: one with my sample obtained from using a kit (RNAqueous kit from Ambion), and the other one, obtained by using the standard protocol with trizol.
My problem is that I could see both bands from rRNA's subunits in the 1st gel, but only one band in the 2nd one. Don't know what that only band would be for, and why there aren't 2!
I obtain much more yield with the 2nd method, so it's the one I need to develop since I need a pretty big amount of RNA for sequenciation.

Hope you can help me! Thanks in advanced!

#2 Trof

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Posted 26 September 2012 - 11:18 AM

Maybe a picture would help.

Our country has a serious deficiency in lighthouses. I assume the main reason is that we have no sea.

I never trust anything that can't be doubted.

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#3 Flopilus

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Posted 26 September 2012 - 04:49 PM

Ok, the first one is the one with the kit and the other one with trizol, the samples are different conditions of the same plant.

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#4 Trof

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Posted 26 September 2012 - 11:42 PM

The first mistake is you didn't put any marker to your gel. Now you don't know what size the band is and how well the bands are separated.
I suggest running the gel again (is it denaturing gel, right? what % of agarose?) with a suitable marker and maybe put your kit RNA and Trizol RNA side by side for comparison.
Your bands on right are more diffused than AFAIK they should be, so it would be maybe helpful to run both RNA in same conditions.

Our country has a serious deficiency in lighthouses. I assume the main reason is that we have no sea.

I never trust anything that can't be doubted.

'Normal' is a dryer setting. - Elizabeth Moon






Also tagged with one or more of these keywords: RNA electrophoresis, plant RNA, band resolution

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