DNase treatment of RNA sample

Tracy Aquilla aquilla at salus.med.uvm.edu
Thu Mar 16 10:42:34 EST 1995


In Article <199503131114.UAA28466 at bekkoame.bekkoame.or.jp>,
tishizuk at AQUARIUS.BEKKOAME.OR.JP (Toshiharu Ishizuka) wrote:
>Dear bionetters
>
>        We are doing differential display experiment. During the
>purification of RNA to remove the chromosomal DNA, we have a serious
>trouble. 
>
>        We purified RNA from heart by Chomczynski method (with TRIZOL LS
>Reagent). However, after the DNase treatment of the RNA, most of the RNA
>was degraded. We used the RNase-free DNase I of Boehringer Mannheim. We
>think that the cause of the degradation is the contamination of RNase in
>RNA sample itself or DNase.
>
>        We would like to know whether the RNA sample purified by
>Chomczynski method contains some RNase. If so, is it necessary to treat the
>RNA with some additional method (phenol extraction etc.) ? We also
>appreciate if somebody inform me the supplier's name of good DNase I. Any
>comment is O.K. Please teach me.
>
>Sincerely yours,
>
>Dr. Toshiharu Ishizuka
>Department of Biochemistry, Chiba University, School of Medicine
>1-8-1, Inohana, Chuo-ku, Chiba, 260 JAPAN
>FAX +81-43-226-2041
>E-mail
>QFH03407 at niftyserve.or.jp
>tishizuk at aquarius.bekkoame.or.jp

I do not think there would be any active RNAse in an RNA prep made using the
protocol you mentioned. If so, you would not get any intact RNA from the
prep. I also use RNAse-free DNAse I from Ambion, and I have never had any
problems with it. Their enzyme is extremely pure; one can do a several-fold
overdigestion with it with no degradation of the RNA. Good luck.
    tracy



More information about the Methods mailing list