I want to do an electrophoresis of RNA from baker's yeast. However, our lab only has a vertical electrophoresis, while usually electrophoresis of nucleic acid (RNA or DNA) use horizontal electrophoresis with agarose gel. Is it possible to do electrophoresis of RNA in vertical electrophoresis using acrylamide gel or agarose? and how will be the quality compared with the horizontal electrophoresis?
I saw in previous literature the two major species of RNAA seen in the gels have size about 2200 and 3900 nucleotides.
Thank you.
Hello there,
you can resolve RNA in either Agarose- or Acrylamide-gels, the orientation of the gel chamber does not matter.
In our lab we are interested in small (~75bp) RNAs which can be nicely resolved by a 10% UreaPAGE. Comparable to agarose-gels, you can adjust the percentage of the gel according the fragments you want to resolve (see attached files).
So in this case, you´d need to lower the acrylamide percentage to around 6% i´d guess. (see https://www.thermofisher.com/order/catalog/product/EC6865BOX#/EC6865BOX) In the end you will have to try youself whether agarose or acrylamid will be the best suited for you goals. Both can be used.
Agree with David Scherf, gel orientation does not matter. Large proteins (about 2 000 nucleotides) can be resolved in 8% polyacrylamide gel, so you can use 6 or maybe 7% gel.
David Scherf I'm curious. How do you visualize the results in the case of Urea/PA gels? Do you wash the gel in running buffer containing EtBr or what?
Alejandro Martin, indeed you are right, that´s exactly how we do it.
We use EtBr and SYBR Gold for staining. In this case i used SYBR Gold since it is more sensitive when compared with EtBr, see attachment.
SYBR Gold: https://www.thermofisher.com/order/catalog/product/S11494#/S11494
You can easily see both native and irreversibly denatured by glyoxal RNA molecules in one 1.5% agarose gel.
Good luck!
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