Well Ed can clarify. His original post is somewhat ambiguous: he didn't really say what he wanted, just that he wanted to precipitate the RNA. It sounded to me like he wanted to get rid of RNA from a plasmid miniprep, especially with his follow up post. But I agree that if he wanted high quality RNA, alkaline lysis is not an appropriate method.
Sent from my iPhone On Mar 10, 2013, at 10:10 PM, Hiranya Roychowdhury <[email protected]> wrote: > Looks like you misunderstood the original post. He wants to get RNA from an > alkaline prep. That is not possible. > > > Hiranya S. Roychowdhury, Ph.D. > Associate Professor > Health & Public Services > NMSU-Dona Ana Community College > 575 527 7725 (office) > > ________________________________________ > From: [email protected] > [[email protected]] on behalf of Michael L. Sullivan > [[email protected]] > Sent: Sunday, March 10, 2013 1:12 PM > To: Ed Siefker > Cc: [email protected] > Subject: Re: LiCl precipitation? > > DNase-free RNase, that is! > > On Mar 8, 2013, at 9:55 AM, Ed Siefker <[email protected]> wrote: > >> Spec reading tells me I got about 150ug of nucleic acid, and >> I'm sure that's not all plasmid DNA. Running it on a gel >> shows a little bit of plasmid DNA, and a large bright splotch >> running around 100bp. No genomic DNA is stuck in the >> wells. >> >> I figure that blotch either has to be RNA, or DNA. If my >> lysis was harsh enough that it degraded the DNA to 100bp >> fragments, I would expect to see the small amount of >> plasmid as a smear instead of a well defined band. >> >> On Thu, Mar 7, 2013 at 9:42 PM, Hiranya Roychowdhury <[email protected]> >> wrote: >>> How do you know that you got RNA? Alkaline lysis does not give you globs >>> of RNA. >>> >>> >>> Hiranya S. Roychowdhury, Ph.D. >>> Associate Professor >>> Health & Public Services >>> NMSU-Dona Ana Community College >>> 575 527 7725 (office) >>> >>> ________________________________________ >>> From: [email protected] >>> [[email protected]] on behalf of Ed Siefker >>> [[email protected]] >>> Sent: Thursday, March 07, 2013 2:29 PM >>> To: [email protected] >>> Subject: LiCl precipitation? >>> >>> I did an alkaline lysis miniprep and ended up with gobs and gobs of RNA. >>> I added an equal volume of 4M LiCl, chilled at -20C for 40 minutes, and >>> spun at 16K Gs for 20 minutes. I don't see a pellet. What am I doing >>> wrong? >>> >>> _______________________________________________ >>> Methods mailing list >>> [email protected] >>> http://www.bio.net/biomail/listinfo/methods >>> >>> >>> >>> _______________________________________________ >>> Methods mailing list >>> [email protected] >>> http://www.bio.net/biomail/listinfo/methods >> >> _______________________________________________ >> Methods mailing list >> [email protected] >> http://www.bio.net/biomail/listinfo/methods > > _______________________________________________ > Methods mailing list > [email protected] > http://www.bio.net/biomail/listinfo/methods > > > > _______________________________________________ > Methods mailing list > [email protected] > http://www.bio.net/biomail/listinfo/methods _______________________________________________ Methods mailing list [email protected] http://www.bio.net/biomail/listinfo/methods
