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Isolating DNA and RNA from the same sample


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#1 AnneC

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Posted 02 December 2004 - 02:33 PM

Hi !!

I'm gonna have to isolate DNA and RNA from the samples (mammalian cells) in a couple of days.
For RNA, we use usually the QIAGEN RNAeasy minikit. I was planning to take an aliquot of cells for DNA and carry on with the rst with this kit for RNA, but I'm wondering now if there wouldn't be a solution to extract both RNA and DNA at the same time.
I've realised that QIAGEN have a kit fot that but I won't have time to get it for my experiment. In the contrary, other companies like Ambion don't have such kits and I found a protocol on their website to extract DNA from organic phases and interfaces kept during RNA isolation. Would it be possible to do the same thing with the QIAGEN kit ??
What would you advice me to do ?
Thanks for your answers.

Anne

#2 pcrman

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Posted 02 December 2004 - 06:48 PM

Hi Anne,

If you want to extract RNA and DNA from the same samples you can use TriReagent from http://www.mrcgene.com/

Actually, TriReagent allows you to extract RNA, DNA and protein from the same sample. I use it for routine RNA extraction even if I have no intention to extract DNA and protein, because the procedure is very simple and cheap. However, I have always found that the DNA quantity I can get is not good enough but that is not a problem for me because I usually only need several micrograms of DNA. The original protocol for subsequent DNA extraction was very time-consuming, but the new protocol has improved a lot.

#3 Solomona

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Posted 03 December 2004 - 12:59 AM

Lyse your cells like you need to and extract total nucleic acids via phenol: chloroform and isopropanol precipitation with RNAse free material.
Divide your aliquot in two parts and use one for DNA purification and one for RNA. I tried both, acidious phenol: chloroform extraction and RNAse free DNAse treatment followed by Proteinase K digestion, like Ambion suggests, and both worked well in reverse transcription. Somehow the QIAGEN RNAeasy minikit didn't work with my total nucleic acid mix.

Check your nucleic acid aliquot before continuing with purifications to avoid extra work with scrambled RNA. Clear ribosomal RNA bands indicate quality RNA. When you have a lot of DNA in your sample, you may have to carry out acidious phenol:chloroform two or three times.

I hope that helped.




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