Removing any traces of DNA

John Dixon jpcd0 at mole.bio.cam.ac.uk
Thu Dec 21 06:57:26 EST 1995


In article <aquilla.1169708309O at emba-news.emba.uvm.edu>,
aquilla at salus.med.uvm.edu (Tracy Aquilla) wrote:

> >On Thu, 14 Dec 1995, Jean Peluso wrote:
> >
> >> I would like to know how I can remove completely DNA from a preparation
> >> of in vitro synthetised RNA. I have already tried the DNAse I recommended
> >> by Invitrogen (T3 Transcription Kit), but when I perform RT-PCR on the
> >> RNA  preparation I unmask traces of DNA, even by using more DNAse and

snip

> has RT activity? I routinely use RNAse-free DNAse I from Ambion to eliminate
> contaminating DNA from my RNA preps and haven't had any problems eliminating
> the contaminating DNA. This enzyme is so clean you can do a several-fold
> over-digest without affecting the RNA. Have you tested the effectiveness of
> your DNAse solution?
>     Tracy



How do you guys then get rid of the DNAse1 prior to cDNA synthesis? If a
phenol chloroform extraction is used, must care be taken to use RNA phenol
ie low pH phenol to avoid loss of the RNA to the wrong phase?

-- 
John Dixon                                        Lab 44 (1223) 334131
Wellcome/CRC Institute                            Fax 44 (1223) 334134
Department of Genetics
Cambridge University    
United Kingdom                           e-m: jpcd0 at mole.bio.cam.ac.uk



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