From BIOSCI-REQUEST Mon May 2 06:10:22 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id GAA27334 for rna-list; Mon, 2 May 1994 06:10:22 -0700 Received: from neutron.chem.yale.edu (NEUTRON.CHEM.YALE.EDU [130.132.25.75]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id GAA27331 for ; Mon, 2 May 1994 06:10:20 -0700 Received: by neutron.chem.yale.edu (931110.SGI.ANONFTP/931108.SGI.AUTO.ANONFTP) for rna@net.bio.net id AA03166; Mon, 2 May 94 09:10:59 -0400 From: eskay@neutron.chem.yale.edu (Sarah Keill) Message-Id: <9405021310.AA03166@neutron.chem.yale.edu> Subject: abortive txns To: rna@net.bio.net Date: Mon, 2 May 1994 09:10:58 -0400 (EDT) X-Mailer: ELM [version 2.4 PL23] Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit Content-Length: 1733 > I am having a problem with aboritve transcription. The buffer I am > using contains 40mM tris pH 8.1, 42C, 5mM DTT, 1mM Spermidine, 22 mM MgCl2, > 50 ug/ml BSA, 80mg/ml PEG 8000, and 0.01% tritonX-100. The template is a > single stranded 70 base DNA with only the region of the promoter ds. This > is at a conc. of 4 uM. 4 mM in each NTP and 15,000 U of T7RNAP are used in > 500 ul. Incubation is 4 hrs. at 42C. > With other templates this works great, but with this particular one I > get virtually no full length product. It has one predicted region of > secondary structure that involves the first three bases of the transcript > with bases 47-49 of the transcript. given the length of your transcript and the potential for secondary structure, an easy first step would be to try the fully double stranded template approach. polymerase should be able to melt secondary structure out, i think, but it doesn't hurt to give it a hand. using a double-stranded template would be more easily done within the context of plasmid-directed reactions. there are easy- to-use puc-derived vectors with t7-necessary equipment available. in my department, it has become almost matter of course to do txns of such long pieces with plasmid templates. additional suggestions: play with the temperature and the dtt concentration. t7rnap is very unstable even at 37, and i would wager you have little active enzyme left at the end of 4hrs at 42. in my lab, people have increased dtt to as high as 100mM to keep the polymerase in the right oxidation state for as long as possible. it doesn't hurt anything and a more stable polymerase is a more processive polymerase. good luck! sarah keill yale university, mb&b and chem From BIOSCI-REQUEST Mon May 2 11:44:00 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id LAA13945 for rna-list; Mon, 2 May 1994 11:44:00 -0700 Received: from HAL.HAHNEMANN.EDU (hal.hahnemann.edu [192.54.238.37]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id LAA13942 for ; Mon, 2 May 1994 11:43:56 -0700 From: springerj@hal.hahnemann.edu Received: by hal.hahnemann.edu (MX V4.0 VAX) id 9; Mon, 02 May 1994 14:45:01 EST Date: Mon, 02 May 1994 14:45:00 EST To: rna@net.bio.net Message-ID: <0097DD52.4B60C940.9@hal.hahnemann.edu> Subject: Does anyone out there know of a source of the FAS/Apo-1 cDNA? I am interested in studying mRNA expression levels in amyotrophic lateral sclerosis. I am also looking for a good human monoclonal as well. Thanks. Joe E. Springer. From BIOSCI-REQUEST Tue May 3 09:11:09 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id JAA20902 for rna-list; Tue, 3 May 1994 09:11:09 -0700 Received: from east.gsfc.nasa.gov (east.gsfc.nasa.gov [128.183.104.4]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id JAA20899 for ; Tue, 3 May 1994 09:11:05 -0700 From: peris@biovx1.dnet.nasa.gov Received: from BIOVX1.DECnet MAIL11D_V3 by east.gsfc.nasa.gov (5.57/Ultrix3.0-C) id AA03882; Tue, 3 May 94 12:08:40 -0400 Date: Tue, 3 May 94 12:08:40 -0400 Message-Id: <9405031608.AA03882@east.gsfc.nasa.gov> To: "rna@net.bio.net"@EAST.DNET.NASA.GOV Subject: double stranded RNAse... I am interested in finding an enzyme that is specific for double stranded RNA, (with no effect on single stranded RNA...was that a redudant statement?) Does anybody know where I can find such a beast comercially.... Thank you Marian Peris From BIOSCI-REQUEST Tue May 3 10:15:18 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id KAA24263 for rna-list; Tue, 3 May 1994 10:15:18 -0700 Received: from dartvax.dartmouth.edu (dartvax.dartmouth.edu [129.170.16.4]) by net.bio.net (8.6.8.1/8.6.6) with ESMTP id KAA24250 for ; Tue, 3 May 1994 10:15:15 -0700 Received: from prancer.Dartmouth.EDU by dartvax.dartmouth.edu (8.6.8.1+DND/4.5HUB) id NAA11721; Tue, 3 May 1994 13:15:54 -0400 Message-id: <10732482@prancer.Dartmouth.EDU> Date: 03 May 94 13:15:50 EDT From: Robert.H.Gross@dartmouth.edu (Robert H. Gross) Subject: Re: double stranded RNAse... To: rna@net.bio.net Marian- You might try using Cobra Venom RNAase (sometimes called V1 RNAase). I think it is available from Boehringer. RNAase III also is double strand specific, but it needs to recognize a specific structure (hairpin with bulge). Bob Gross From BIOSCI-REQUEST Thu May 5 08:42:57 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id IAA04147 for rna-list; Thu, 5 May 1994 08:42:57 -0700 Received: from mserv.rug.ac.be (mserv.rug.ac.be [157.193.40.37]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id IAA04144 for ; Thu, 5 May 1994 08:42:52 -0700 Received: from allserv.rug.ac.be by mserv.rug.ac.be with SMTP id AA09120 (5.67b/IDA-1.5 for ); Thu, 5 May 1994 17:42:51 +0200 Received: by allserv.rug.ac.be (5.0/SMI-SVR4) id AA07501; Thu, 5 May 94 17:39:59 +0200 X-Nupop-Charset: English Date: Thu, 5 May 1994 16:43:10 +0100 (MET) From: "Dirk Vandenberghe" Sender: Dirk.Vandenberghe@rug.ac.be Message-Id: <60192.dvdbergh@allserv> To: rna@net.bio.net Subject: INTERNATIONAL PHARMACEUTICAL BIOTECHNOLOGY CONFERENCE Content-Length: 2279 **************************************************************************** *** The 2nd International Pharmaceutical Biotechnology Conference *** **************************************************************************** APRIL, 23 - 27 1995 GHENT, BELGIUM, EUROPE INVITED SPEAKERS (unlimited list 05-05-94) : T. Blundell (UK) H. Galjaard (NL) M. Moereels (Belgium) G. Bleecker (USA) D. Crommelin (NL) M. Mareel (Belgium) D. Brizner (USA) W. Gibbons (UK) M. Soria (Italy) A. Angiuoli (USA) J. De Muth (USA) T. Osslund (USA) N. Burnette (USA) L. Hansen (USA) G. Reeder (USA) R. Braeckman (USA) W. Fiers (Belgium) H. Robays (Belgium) S. Cohen (USA) I. Hart (UK) M. Van Montagu (Belgium) J. Chamberlain (USA) D. Heyd (Israel) K. Seamon (USA) R. Dobbelaere (Belgium) Ph. Johnson (USA) P. Gantz (USA) Y. Yamada (Japan) N. Spurr (UK) J. Cook (USA) S. Huber (USA) President: Prof. Dr. E. Van den Eeckhout University of Ghent FFW Harelbekestraat 72 Tel: 32 9 221 99 43 B-9000 GENT Fax: 32 9 220 66 88 BELGIUM, EUROPE Main topics : Gene Therapy, Vaccines and Monoclonal Antibodies, Drug Delivery Systems, Modification of Plants, Drug Design, Biotechnology in the Treatment of Cancer, Pharmacoeconomics, Human Genome Project, ...... !!! Send your mailing (!) address to Dirk.Vandenberghe@rug.ac.be if you want your name on our mailing list for future information and registration forms, and let me know if you are interested in : # poster session / abstract. # joining the International Association for Pharmaceutical Biotechnology. # exhibitor space (industrial forum). # pre / post - conference tours to beautiful cities in Europe. Dirk.Vandenberghe@rug.ac.be University of Ghent Laboratory Pharmaceutical Biotechnology From BIOSCI-REQUEST Fri May 13 07:54:27 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id HAA28353 for rna-list; Fri, 13 May 1994 07:54:27 -0700 Received: from ania.path.ox.ac.uk (ania.path.ox.ac.uk [163.1.19.2]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id HAA28350 for ; Fri, 13 May 1994 07:54:23 -0700 From: birney@molbiol.ox.ac.uk Received: by molbiol.ox.ac.uk (MX V4.0-1 AXP) id 10; Fri, 13 May 1994 15:54:45 +0100 Date: Fri, 13 May 1994 15:54:43 0000 To: rna@net.bio.net Message-ID: <0097E600.DAF2E911.10@molbiol.ox.ac.uk> Subject: hnrnp prorteins RNA-netters This is another bleated attempt to find anyone with a good idea about nomenculture and general order of the hnrnp proteins and chloroplast RNA binding proteins. I'm maintaining a very small scale database for the RRM or RBD or RNP-CS domain (take your pick for the name) which now has grown somewhat. I'm trying to have an overhaul at the moment to make more of the work automatic. In addition I've moved to searching DNA databases directly with profiles which have allowed me to find virtually all RRM sequences possible to detect. However these two families, hnrnp a1 like proteins and chloroplast RNA binding proteins are enough to give anyone a headache. I need help from people who are actually working (experimentally) with these little blighters to sort out what is what and which sequences should be grouped under which others. So...PLEASE...if you know what is going on please email me. If you know someone else who might know what is going on please persuade him/her to email me. The database is available as a table on a file server. email RRM@molbiol.ox.ac.uk with "send rrm" in the message. Unforunately the PROM database system is down (and will be for some time) as we migrate to a different system and I recode things... any help will be much appreciated ewan birney birney@molbiol.ox.ac.uk From BIOSCI-REQUEST Fri May 13 09:04:48 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.8.1/8.6.6) id JAA01687 for rna-list; Fri, 13 May 1994 09:04:48 -0700 Received: from cray.com (root@timbuk.cray.com [128.162.19.7]) by net.bio.net (8.6.8.1/8.6.6) with SMTP id JAA01684 for ; Fri, 13 May 1994 09:04:46 -0700 Received: from shamu (shamu.cray.com) by cray.com (Bob mailer 1.2) id AA19257; Fri, 13 May 94 11:05:17 CDT Received: by shamu id AA29556; 4.1/CRI-5.4; Fri, 13 May 94 10:35:19 CDT From: mwd@carina.cray.com (Mark Dalton) Message-Id: <9405131535.AA29556@shamu> Subject: Re: Pseudo Knot summary To: rna@net.bio.net Date: Fri, 13 May 1994 10:35:18 -0500 (CDT) X-Mailer: ELM [version 2.4 PL23] Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit Content-Length: 4124 Hi! I posted this awhile ago. I currently use Dr. Michael Zukers code for folding and Don Gilberts for viewing on a Mac. Thanks! Mark Here is what I have heard: >From mcpheeters@biochemistry.cwru.edu Wed Apr 13 19:26:04 1994 Subject: Re: RNA Pseudo-knots In message <9404132235.AA09191@calamity> Mark Dalton writes: > > I am wondering if there is any software available for figuring > out Pseudo-knots. There is a researcher at the Univ. of MN, I am > trying to help out. I don't seem to have much information on this. > > Any information would be appreciated, I will post a summary to the > group. > > Thanks! > > Mark > -- Mark, I have of a couple of references that may describe software that deals with pseudoknots, these are: 1) Prediction of RNA secondary structure, including pseudoknotting, by computer simulation . Abrahams,J.P.,van den Berg,M.,van Batenburg,E., and Pleij,C.W.A. NAR(1990)18,3035-3044 2) RNA pseudoknots: Structure, detection, and prediction Pleij,C.W.A. and Bosch,L. Meth.Enz.(1989)180,289-303 3)Detecting pseudoknots and other local base-pairing structures in RNA sequences Martinez,H.M. Meth.Enz.(1990)183,306-317 Good luck, David McPheeters ------------------------------------------------------------- >From barry@aaRS.mit.edu Wed Apr 13 21:24:58 1994 Subject: RE: RNA Pseudo Knots Hi Mark- I have been working on pseudoknots lately, trying to them into some unusual structures in hopes of gaining some additional stacking stabilization. I have been using MCSYM available from NCBI. It is useful for evaluating the stereochemical feasability of a fold. You do have to write scripts containing the desired structural features so there is a built in "bias". This makes it a rather questionable tool for exhaustive structural searches. You may also consider a simple secondary structure program like Zuker's MFOLD. You can get info on MCSYM from: ncbi.nlm.nih.gov in the pub/MC-SYM directory. You will have to request access to the executables and there are instructions there. MFOLD can be obtained by gopher at the IUBIO archives. I hope this helps. If you have any other thoughts on the subject or get any really insightful suggestions from your request I would be very interested in hearing them. Good luck Barry >From stormo@beagle.Colorado.EDU Thu Apr 14 09:49:25 1994 Subject: RNA Pseudo-knots It depends a little on what type of data you have. If you have a single sequence and want to predict its structure, allowing for pseudoknots, you should look at the paper by abrahams et al, NAR 18:3035. If you have lots of sequences and want to determine the structure that they have in common you can use comparative methods, which allow pseudoknots, such as described in Gutell et a. NAR 20:5785. Gary Stormo >From jweiland@badlands.NoDak.edu Sat Apr 16 09:47:00 1994 Subject: Pseudoknot programs Hi from the Fargo area! Although there are probably RNA folding programs created in North America which have pseudoknot recognition algorithms incorporated into them by now, there may be a free-ware or share-ware version of a program developed by C.W.A Pleij at the University of Leiden (Netherlands). He was the original founder of pseudoknots, and shortly thereafter collaborated with some computer jocks in creating a program for thier recognition. I don't know whis E-mail address, but he should be a memeber of several biophysical chemistry societies. Good luck! John Weiland jweiland@badlands.nodak.edu -- Mark Dalton AUG-GCU-AGA-AAG H Cray Research, Inc. M A R K | Eagan, MN 55121 CH3-S-CH2-CH2-C-COOH Internet: mwd@cray.com | (612)683-3035 NH2 Marks Info/Resume Thoughts to ponder: Life is Paradox: one = many, many = one. Most difficult tasks are easy, it just depends on if you know the solution. All tasks are easy, it all depends on who you ask. From BIOSCI-REQUEST Thu May 26 09:18:53 1994 Return-Path: BIOSCI-REQUEST Received: (from daemon@localhost) by net.bio.net (8.6.9/8.6.6) id JAA12119 for rna-list; Thu, 26 May 1994 09:18:53 -0700 Received: from mailhost.lanl.gov (mailhost.lanl.gov [128.165.3.12]) by net.bio.net (8.6.9/8.6.6) with ESMTP id JAA12116 for ; Thu, 26 May 1994 09:18:51 -0700 Received: from temin.lanl.gov by mailhost.lanl.gov (8.6.8.1/1.2) id KAA11245; Thu, 26 May 1994 10:19:30 -0600 Received: from namot.t-10 (namot.lanl.gov) by temin.lanl.gov (4.1/SMI-4.1) id AA06099; Thu, 26 May 94 10:19:27 MDT Received: by namot.t-10 (5.0/SMI-SVR4) id AA04093; Thu, 26 May 1994 10:20:51 +0700 From: esc@temin.LANL.GOV (Gene Carter) Message-Id: <9405261620.AA04093@namot.t-10> Subject: Nucleic Acid MOdeling Tool To: rna@net.bio.net Date: Thu, 26 May 1994 10:20:50 -0600 (MDT) X-Mailer: ELM [version 2.4 PL23] Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit Content-Length: 4616 Announcing the release of Namot. NAMOT(Nucleic Acid MOdeling Tool) is a tool developed at Los Alamos National Laboratory for the structural manipulation of single or double stranded, RNA or DNA. NAMOT uses a set of reduced co-ordinates to describe base/unit, sugar and phosphate geometry. Detailed description of these parameters can be found in the references listed at the bottom of this announcement. Namot features an X11 graphics interface for display of the molecule while modifications are being carried out. Both Motif and XView toolkit versions exist, which should allow namot to be easily ported to most unix machines. An article describing namot is currently in press(CABIOS,1994). Namot features: Input: PDB,Amber, or parameters Output: Amber, PDB, or parameters, encapsulated postscript. General: Can be used to manipulate ss or ds RNA or DNA via reduced parameters. Which have intuitive meaning, and allow easy manipulation of the base/unit without bond breakage. Auto-adjusts: O3'-P-O5' to allow the phosphate to cross varying gaps. Monitors distances: A simple point and click, connects two atoms with a dashed line and displays the distances whenever the structure is changed. Generation of structures: Generates the parameters for A,B,& Z. RNA/DNA, ss or ds Can be used as a starting place. Label BP: Displays the unit number on the right side of the screen at the same level as the C3' Online help: There is an extensive online help system. Rotations: Easily accessible rotational controls. Currently there are three versions of namot. NAMOT1: The original, uses the XView toolkit and basic monochrome Xlib graphics. It does feature a WYSIWYG(What You See Is What You Get) printing screen that produces encapsulated postscript output. NAMOT 1.2: Features color graphics options, both wireframe and CPK representations. It produces color encapsulated postscript output. A command line lets the user set foreground/background to white or black. The user can set the mode to mono or color(color is the default). The command line also allows the user to query for atom position, angles, and torsional angles. The online help for "commands" lists the specific syntax. NAMOT 1.2s: Same basic features as NAMOT 1.2, but uses the motif toolkit to build the interface and has been tested on an SGI Iris Indigo. Namot is availible over the internet via anonymous ftp. To get namot: ftp namot.lanl.gov login: anonymous Password:(Your email address) cd pub/namot get Note: SGI users may need to use "nlist" and not "ls" to list files. NAMOT 1. and NAMOT 1.2 have been tested on: Sun SparcStation 1+ running SunOS 4.1.3 Sun SparcStation LX running SunOS 2,3 Sun SparcClassic running SunOS 2,2 NAMOT 1.2s has been tested on SGI Iris Indigo References: Base/Unit description: Journal of Biomolecular Structure and Dynamics. Volume 6, Issue Number 3 (1988), page 397 Soumpasis, D.M., and Tung, C.-S. Sugar description: Journal of Biomolecular Structure and Dynamics, Volume 5, Issue 3(1987), page 513 Garcia A.E., and Krumhansl, J.A. Phosphate description: Computation of Biomolecular Structures: Achievements, Problems, and Perspectives, Tung, C-S, Soumpasis, D. M. & Jovin, T. M., eds, Springer-Verlag, NY Feedback on NAMOT is greatly appreciated, email can be sent to namot@temin.lanl.gov. Gene Carter esc@temin.lanl.gov Chang-Shung Tung cst@temin.lanl.gov Copyright 1994 The Regents of the University of California. This software was produced under U.S. Government contract(W-7405-ENG-36) by Los Alamos National Laboratory, which is operated by the University of California for the U.S. Separtment of Energy. The U.S Government is licensed to use, reproduce, and distribute this software. Permission is granted to the public to copy and use this software without charge, provided this Notice and any statement of authorship are reproduced on all copies. Neither the Government nor the University makes any warranty, expressly or implied, or assumes any liability or responsibility for the use of this software. Los Alamos Computer Code:94-6 --------------------------------------------------------------------------- - email:esc@temin.lanl.gov |Gene Carter | - talk: esc@namot.lanl.gov |MS k710 | - Work Phone: 505 662 2591 |Los Alamos National Laboratory | - Group t-10 LANL |Los Alamos, NM 87545 | ===========================================================================