From owner-celegans@net.bio.net Fri Feb 23 22:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!newsfeed.internetmci.com!usenet.eel.ufl.edu!news.ultranet.com!bigboote.WPI.EDU!transfer.stratus.com!news3.near.net!yale!yale.edu!news.ycc.yale.edu!news From: RBURDINE@biomed.med.yale.edu (R.O.S.B.F.) Newsgroups: bionet.celegans Subject: Re: Glass-bead RNA extraction Date: 23 Feb 1996 21:48:15 GMT Organization: Keeper of Dakota, Cat from Hell Lines: 26 Distribution: world Message-ID: <4glcmv$2k9@news.ycc.yale.edu> References: NNTP-Posting-Host: biomed.med.yale.edu X-News-Reader: VMS NEWS 1.20 In-Reply-To: Morris_maduro@biology.ualberta.ca's message of 23 Feb 1996 01:57:47 GMT In Morris_maduro@biology.ualberta.ca writes: > > I have been trying very hard, to no avail, to get RNA from nematodes using > the 'glass-bead' protocol found in WBG Nov '89 p.154. It is a standard > guanidine thiocyanate/organic extraction protocol. I have routinely used the TRIZOL reagent from Gibco-BRL to extract RNA from worms. I have successfully used this RNA in Northen analysis, RT-PCR and RACE experiments. TRIZOL is a reagent that contains the guanidine and the phenol. You solublize thw worms in this solution and add chloroform to separate RNA into the aqueous,DNA at the interface, and proteins in the organic phase. You can, in theory, get all the components back from each of the phases but I have never tried this. The TRIZOL and vortexing is all I have used to solublize the worms. I did not need to use glass beads. One advantage to using this is that the acid pH of the phenol is controlled for by Gibco. It is basically the modified Chomczynski and Sacchi you are currently doing. But it has worked well for me and for other people in the lab. I'd be happy to send you a detailed protocol if you are interested. Becky .