RNA vs DNA purifucation - how are they separated from each other? (Jun/23/2005 )

Hi,
Maybe this is not a good question but if someone can answer me i will really appreciate it.
During the DNA and RNA purification it is only the use of basic or acid phenol the main point for them to be separated from each other. I mean, when you get the electrophoresis results after purifying, how can you be sure that what you are looking at the picture is ARN and not DNA. In order to purify nucleic acids you are using basically the same substances (Ethanol, phenol, chloroform,even the the same extraction buffer can be used for both).
If it is the phenol, how does it work?

Thanks in advance

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hi
ph for rna extraction is about 4.7 and for DNA it's a mix phenol chloroform isoamylalcohol whithout adjusting ph. Hence i'm not sure about the pH of this prep.
it's functionning by exclusion of the non-wanted type...
but i'm not a chemist

hope that helps.
fred

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Errr, well, that's totally new to me, we always adjusted the pH of our phenol-
solution to 7.5 by equilibration with TE.
Phenol alone is an acid, the pH will be quite low without adjusting the pH.
This will lead to protonation of the ribose-phosphate backbone of the DNA and
partitioning of the DNA into the phenol-phase, therefore you will receive lower
yields.
Also the commercially available phenol-solutions are adjusted to pH 7.5 or above.

Regards,
Hennry

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thanks for the input. That's why i never adjust ph for DNA. Its a pre adjusted ph...

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you could take a sample and add RNase and them run an electrophoresis... if you see the band... that's DNA.

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Genomic DNA will run as a single band at about 23kd. With perhaps some degradation as a smear.

RNA will run two main bands 28S and 18S with the 5S further down and much fainter. Any degradation will be a big blob of white at the bottom of the gel.

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