From owner-rapd@net.bio.net Fri Mar 01 22:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!elroy.jpl.nasa.gov!swrinde!howland.reston.ans.net!vixen.cso.uiuc.edu!usenet.ucs.indiana.edu!willow.bio.indiana.edu!user From: dewolf@bio.indiana.edu (Diana Wolf) Newsgroups: bionet.molbio.rapd Subject: Re: Thermostability of Taq polymerase Date: 2 Mar 1996 03:56:01 GMT Organization: Indiana University Lines: 24 Distribution: world Message-ID: References: <4gtse8$3lt8@yuma.ACNS.ColoState.EDU> NNTP-Posting-Host: willow.bio.indiana.edu In article , abartlet@ASRR.ARSUSDA.GOV ("Alan C. Bartlett") wrote: > I don't know the maximum temperature, but most experimenters use 94 - > 95 C as the maximum. Maybe you are putting too much template DNA in > your reaction mixture. I have found that to be critical in my work. > Try a series of dilutions of your typical template solution and see if > you can get amplification. Do you know why one gets better amplification with less template? I find that less template is better also, and the same is true when I use microsatelite primers. It seems counterintuitive to me. We store our genomic DNAs in TE (tris and EDTA), and I have always wondered if the increased reaction with lower amounts of template was actually because we were putting less EDTA in the reaction rather than because we were putting less template in the reaction. Diana -- Diana Wolf Biology Department Indiana University From owner-rapd@net.bio.net Sat Mar 02 22:00:00 1996 Path: biosci!daresbury!not-for-mail From: chn Newsgroups: bionet.molbio.rapd Subject: subscription sought Date: 3 Mar 1996 09:10:38 -0000 Lines: 7 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <4hbnme$su1@mserv1.dl.ac.uk> Original-To: farmer@yvax.byu.edu request subscription for the biosci newsgroup on rapd. please forward info,message,subscription,any other relevent details to chndan@mcbl.iisc.ernet.in e-mail is the only mode available. thanks. From owner-rapd@net.bio.net Sun Mar 03 22:00:00 1996 Path: biosci!plantpath.ksu.edu!kzeller From: kzeller@plantpath.ksu.edu ("Kurt Zeller") Newsgroups: bionet.molbio.rapd Subject: Re: Thermostability of Taq polymerase Date: 3 Mar 1996 19:41:43 -0800 Organization: K.S.U. Plant Pathology Lines: 39 Sender: daemon@net.bio.net Distribution: world Message-ID: <552A2914F5A@plantpath.pp.ksu.edu> NNTP-Posting-Host: net.bio.net > In article , > abartlet@ASRR.ARSUSDA.GOV ("Alan C. Bartlett") wrote: > > > I don't know the maximum temperature, but most experimenters use 94 - > > 95 C as the maximum. Maybe you are putting too much template DNA in > > your reaction mixture. I have found that to be critical in my work. > > Try a series of dilutions of your typical template solution and see if > > you can get amplification. > > > Do you know why one gets better amplification with less template? I find > that less template is better also, and the same is true when I use > microsatelite primers. It seems counterintuitive to me. We store our > genomic DNAs in TE (tris and EDTA), and I have always wondered if the > increased reaction with lower amounts of template was actually because we > were putting less EDTA in the reaction rather than because we were putting > less template in the reaction. > > Diana As I understand it one of the benefits of having less template DNA in the reaction is that your primers have less competition for binding sites. More dilute template DNA reduces the chance of the binding sites finding a homolog that is not your primers. Hope this helps a bit Kurt ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ Kurt Zeller Throckmorton Hall Dept. of Plant Pathology Kansas State University Manhattan, KS 66506 "Mock not the procrastinators,... for they will be the last to die!" From owner-rapd@net.bio.net Sun Mar 03 22:00:00 1996 Path: biosci!ASRR.ARSUSDA.GOV!abartlet From: abartlet@ASRR.ARSUSDA.GOV ("Alan C. Bartlett") Newsgroups: bionet.molbio.rapd Subject: Re: Thermostability of Taq polymerase Date: 4 Mar 1996 12:44:04 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 45 Sender: daemon@net.bio.net Distribution: world Message-ID: References: NNTP-Posting-Host: net.bio.net I do not store my DNA in TE so I guess that EDTA is not the limiting factor (at least in my reactions). I have postulated (silently to myself) that too much DNA ties up the primers so that matched sites are not made and that there is therefore no double stranded DNA to amplify. I don't know if that is possible, but it satisfies my curiosity so I dilute my DNA and go on about my business. I think that I might even get by on less template than I now use, because every once in a while I still get a blank lane when the florometer shows DNA is present. On 2 Mar 1996, Diana Wolf wrote: > In article , > abartlet@ASRR.ARSUSDA.GOV ("Alan C. Bartlett") wrote: > > > I don't know the maximum temperature, but most experimenters use 94 - > > 95 C as the maximum. Maybe you are putting too much template DNA in > > your reaction mixture. I have found that to be critical in my work. > > Try a series of dilutions of your typical template solution and see if > > you can get amplification. > > > Do you know why one gets better amplification with less template? I find > that less template is better also, and the same is true when I use > microsatelite primers. It seems counterintuitive to me. We store our > genomic DNAs in TE (tris and EDTA), and I have always wondered if the > increased reaction with lower amounts of template was actually because we > were putting less EDTA in the reaction rather than because we were putting > less template in the reaction. > > Diana > > -- > Diana Wolf > Biology Department > Indiana University > > Alan C. Bartlett "GENES-R-US" "ALTERATIONS AVAILABLE!^" "^some restrictions may apply:)" From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!ns1.faseb.org!lamarck.sura.net!newsfeed.internetmci.com!in1.uu.net!newsflash.concordia.ca!feed.umontreal.ca!alize.ERE.UMontreal.CA!landryp From: landryp@ERE.UMontreal.CA (Landry Pierre-Alexandre) Newsgroups: bionet.molbio.rapd Subject: Re: Life expectancy of Taq Date: 5 Mar 96 22:27:42 GMT Organization: Universite de Montreal Lines: 12 Distribution: world Message-ID: References: <4hgghg$40hk@yuma.ACNS.ColoState.EDU> NNTP-Posting-Host: alize.ere.umontreal.ca dpicton@lamar.colostate.edu (Deric Picton) writes: >What is the life expectancy of Taq once it has reached its >expiration date provided by the manufacturer? My Taq is at its >expiration date and need to know if it should be good. > Deric Picton I have been using a full tube of TAQ that was expired for 6 months and still got an amplification product. As long as it was kept in the right conditions, I figure that TAQ can last longer than the manufacturer suggests. And anyway, "best before" doesn't mean "no good after" ;) From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!daresbury!not-for-mail From: Abd elnasser Elashry Newsgroups: bionet.molbio.rapd Subject: more information Date: 5 Mar 1996 16:51:18 -0000 Lines: 18 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <4hhre6$1ct@mserv1.dl.ac.uk> Original-To: rapd@dl.ac.uk Hello for Everybody: I am a new subscriber in your discussion group. I am a master student my study is about Molecular characterization of Meloidogyne spp. I knew that I can use sonic dismembrator as a tool to break out the cuticle of juveniles, but I don't know more details. the information about that will help me to extract DNA as a template in PCR reactions. So, it will be very useful if anyone could send me more details about this method, or any information about the references which is belong to this point. Thanks for your time, and your cooperation. Yours Abd Elnasser ************************************************************* Abd El-nasser Mohamed Abd El-Moneim Elashry Agricultural Research Center (ARC). Agricultural Genetic Engineering Research Institute (AGERI). 9 Gamaa St., Giza 12619 Egypt ************************************************************* From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!ACD.TUSK.EDU!Prakash From: Prakash@ACD.TUSK.EDU (C. S. Prakash) Newsgroups: bionet.molbio.rapd Subject: News Report in Biotechnology Date: 5 Mar 1996 12:45:06 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 31 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net A monthly newsletter "NBIAP News Report" that describes briefly the news and research items of interest to agricultural biotech community is available both in print and electronic version. For internet access to the News Report, textfiles, and databases use one of the following procedures. 1. Through WWW: http://www.nbiap.vt.edu/ 2. Use telnet or gopher to connect to ftp.nbiap.vt.edu 3. Use ftp to connect to ftp.nbiap.vt.edu. Use "anonymous" as your user-id, your email address as your password. Type "cd pub/nbiap". To have the News Report automatically emailed, send an email message to news@nbiap.biochem.vt.edu and type SUBSCRIBE NEWSREPORT in the message section. National Biological Impact Assessment Program (NBIAP) runs this Information Systems for Biotechnology, which is a joint project of USDA/CSREES and the Virginia Polytechnic Institute and State University. It does not necessarily reflect the views of the U.S. Department of Agriculture or of Virginia Tech. The News Report may be freely photocopied or otherwise distributed without charge. . For Additional Information: P.L. Traynor, Editor, Information Systems for Biotechnology, 120 Engel Hall, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0308, tel: 540-231-2620, fax: 540-231-2614, email: nbiap@vt.edu From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!COMP.UARK.EDU!jamiles From: jamiles@COMP.UARK.EDU (jay miles) Newsgroups: bionet.molbio.rapd Subject: rapds in cotton Date: 5 Mar 1996 07:51:04 -0800 Organization: U of AR Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: <313C6B19.1F9C@comp.uark.edu> NNTP-Posting-Host: net.bio.net I have a friend interested in doing rapd research with cotton. If anyone knows of any useful protocols or primers, I'm sure he would be interested in hearing from you. Contact Altaf Khan at makhan@comp.uark.edu ! Thanks From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!COMP.UARK.EDU!jamiles From: jamiles@COMP.UARK.EDU (jay miles) Newsgroups: bionet.molbio.rapd Subject: test Date: 5 Mar 1996 07:39:48 -0800 Organization: U of AR Lines: 1 Sender: daemon@net.bio.net Distribution: world Message-ID: <313C6879.76EA@comp.uark.edu> NNTP-Posting-Host: net.bio.net just a test post! From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!daresbury!nntp-trd.UNINETT.no!newsfeed.sunet.se!news00.sunet.se!sunic!mn6.swip.net!plug.news.pipex.net!pipex!tube.news.pipex.net!pipex!lade.news.pipex.net!pipex!tank.news.pipex.net!pipex!usenet.eel.ufl.edu!news.bright.net!chi-news.cic.net!nntp.coast.net!col.hp.com!csn!news-1.csn.net!csn!nntp-xfer-2.csn.net!yuma!usenet From: dpicton@lamar.colostate.edu (Deric Picton) Newsgroups: bionet.molbio.rapd Subject: Life expectancy of Taq Date: 5 Mar 1996 04:39:12 GMT Organization: Colorado State University, Fort Collins, CO 80523 Lines: 4 Message-ID: <4hgghg$40hk@yuma.ACNS.ColoState.EDU> NNTP-Posting-Host: ts2209.slip.colostate.edu X-Newsreader: SPRY News 3.03 (SPRY, Inc.) What is the life expectancy of Taq once it has reached its expiration date provided by the manufacturer? My Taq is at its expiration date and need to know if it should be good. Deric Picton From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!CS.Arizona.EDU!news.Arizona.EDU!hamblin.math.byu.edu!acs2.byu.edu!news.cuny.edu!caen!newsxfer2.itd.umich.edu!chi-news.cic.net!nntp.coast.net!fu-berlin.de!zrz.TU-Berlin.DE!cs.tu-berlin.de!uni-erlangen.de!news.th-darmstadt.de!news.uni-mainz.de!usenet From: Quicksilver Newsgroups: bionet.molbio.rapd Subject: WAF1 Date: Tue, 05 Mar 1996 14:42:27 +0000 Organization: Johannes Gutenberg-Universität Mainz, Germany Lines: 2 Message-ID: <313C52D3.46E8@vzdmzd.zdv.uni-mainz.de> NNTP-Posting-Host: atobec.toxikologie.uni-mainz.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Macintosh; I; 68K) Who knows something about the genomic organization and intron sequences of p21WAF1 (CIP1)? From owner-rapd@net.bio.net Mon Mar 04 22:00:00 1996 Path: biosci!agate!howland.reston.ans.net!swrinde!newsfeed.internetmci.com!in1.uu.net!newsflash.concordia.ca!feed.umontreal.ca!alize.ERE.UMontreal.CA!landryp From: landryp@ERE.UMontreal.CA (Landry Pierre-Alexandre) Newsgroups: bionet.molbio.rapd Subject: why does less template DNA works better Date: 5 Mar 96 22:33:24 GMT Organization: Universite de Montreal Lines: 21 Distribution: world Message-ID: NNTP-Posting-Host: alize.ere.umontreal.ca Hello there! I noticed that this question was asked lately in this group, so I thought that I may bring my own explanation to this problem. The thing I see there is the fact that RAPD usually uses 10-mers primers, so the annealing temperature must be quite low. But the two complementary DNA strands, after denaturation, can anneal at quite high temperature I guess. So if you have a too high concentration of template DNA, the two brands will easily anneal before the primers can hope to do so (since it is a reaction of competition). But if the DNA concentration is low, the probability that the two brands will meet is reduced, and the higher primer concentration will favor primer annealing, and then generate bands. I think the keyword is "competition". Anyways, this is only a suggestion. Pierre-Alexandre Landry Dept. des Sciences Biologiques Universite de Montreal C.P. 6128, succursale centre ville Montreal, Quebec H3C 3J7 From owner-rapd@net.bio.net Tue Mar 05 22:00:00 1996 Path: biosci!daresbury!not-for-mail From: Louis van de Zande Newsgroups: bionet.molbio.rapd Subject: Re: why does less template DNA works better Date: 6 Mar 1996 07:17:16 -0000 Organization: Department of Biology, RUGroningen Lines: 26 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <4hje5s$69t@mserv1.dl.ac.uk> Reply-To: L.P.W.G.M.van.de.Zande@biol.RUG.NL Original-To: rapd@dl.ac.UK > Hello there! > > I noticed that this question was asked lately in this group, so I > thought that I may bring my own explanation to this problem. > The thing I see there is the fact that RAPD usually uses > 10-mers primers, so the annealing temperature must be quite low. But the > two complementary DNA strands, after denaturation, can anneal at quite > high temperature I guess. So if you have a too high concentration of > template DNA, the two brands will easily anneal before the primers can > hope to do so (since it is a reaction of competition). I'm not so sure about that. There is a lot of primer out there. And "single copy" DNA probably takes more time to reanneal than the short period between denaturing and amplification (at the most some 4 min, depending on your cycler and cycle-profile). But certainly competition and complexity of template are important "concerns". L. van de Zande Dept. Genetics Groningen Netherlands From owner-rapd@net.bio.net Tue Mar 05 22:00:00 1996 Path: biosci!fao.org!ICPPGR From: ICPPGR@fao.org (ICPPGR, by way of Prakash@acd.tusk.edu (C. S. Prakash)) Newsgroups: bionet.molbio.rapd Subject: Diversification of Vegetable Crops Date: 6 Mar 1996 11:08:12 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 379 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net SECOND ANNOUNCEMENT Third International Symposium on Diversification of Vegetable Crops Pre-registration deadline July 1.1996 The International Society for Horticultural Science ( Working Group of the Section for Vegetables) and the organizing committee are honored to invite you to participate in the Third International Symposium on Diversification of Vegetable Crops (ISDVC), to be held in September 24 - 27, 1996, in Beijing, China. Post symposium tours are planned to visit different part of China. The Third International Symposium on Diversification of Vegetable Crops provides a forum for the exchange of information on all aspects of vegetable crops, especially on crops not grown commercially in the main production regions. All contributions are welcome if related to the above subject. Emphasis will be given to the following topics: * Germplasm evaluation and utilization; * Genetic resources and breeding; * Crop physiology and ecology; * Growing methods and systems; * Nutritional value and special natural compounds for improvement of health; * Quality standards and post-harvest physiology and technology; * Food utilization and processing and * Economics, marketing and trade. The Symposium will begin with a plenary session of keynote speakers on Tuesday morning September 24 followed by oral presentations on divided programs in the afternoon. A reception will be held in the evening at China - EU Center for Agricultural Technology (CECAT). September 25 will be dedicated to oral papers and posters. A general tour which highlights the local vegetable research institution, production and market will be conducted on September 26. On the following day, this discussion will be continued, then the symposium will be concluded. Call for papers Abstracts Titles and abstracts submitted for consideration must be sent to the Symposium secretariat Mr. Hu Xiaoguang and no later than April 1, 1996. Abstracts should be typed single-spaced on plain paper using the format below: Title: Author(s): Address: Abstract: Preference: Oral or poster The abstract should not exceed 200 words and must emphasize objectives and results. Inclusion of tentative or final conclusions will greatly strengthen abstracts. Authors should indicate their willingness to present a poster in lieu of an oral presentation, as the number of oral presentation is limited by time. Authors will be advised of acceptance or rejection of their abstract and its form of presentation, whether oral or poster, by May 1, 1996. Electronic submission of abstracts is preferred, either via e-mail or on 3.5 diskettes. Posters Posters will be displayed continuously during the Symposium. There is no limit to the number of posters that an author can present, but space limitation will dictate the maximum number of posters accepted for the meeting. Poster space A space 2m x 1m (78UL x 39UL ) is available for each poster. Poster Identification Abstract numbers, titles, names and affiliations should appear on the top of the poster. A simple sans serif-face font, e.g., Helvetica, should be used. Lettering for the abstract number and title should be least 1 inch: The authorsur names and affiliations may be somewhat smaller. Authors are urged to include photographs to assist in author identification. Poster Content Do not prepare a poster as if it is a manuscript. Primarily use tables and figures and limit verbiage. Details of the work can be provided in discussions with interested parties. Lettering for text and illustrations should range in size between 6 mm and 12 mm. Poster Mounting A poster backing will be provided for each poster. Push- pin or Velcro buttons may be used. Pins and Velcro must be supplied by the presenter - they will not be available on-site. Oral Presentations Oral presentations are limited to 15 min. (12-min. talk and 3-min. discussion ). Speakers should pay particular attention to the quality of their visual aids, rehearse their presentation thoroughly, and stay within their allotted time. A 35-mm slide projector and standard overhead projector will be available in each sessionurs meeting room. Authors must bring their own slide carousels. A room to preview slides will be available throughout the meeting. Proceedings Accepted paper is limited to 8 pages, single-spaced. Tables and figures are including within that limit. Instructions for camera-ready manuscript preparation will be sent to authors of accepted abstracts. Completed manuscripts will be due August 1, 1996. All accepted abstracts will be printed and distributed at the Symposium. Language All abstracts, posters, papers, and oral presentations will be in English. Meeting Location and Hotel: The Symposium will be held at the China - EU Center for Agricultural Technology located between the Great Wall Hotel and the Lufthansa Shopping Center in 55, Nongzhang Beilu, Chaoyang District, Beijing China. The room rate with free breakfast will be 40 US$ per night for single or double occupancy. The hotel will provide buffet service which cost around 6 US$ per meal. Transportation Many international air provide frequent service to the Beijing Airport. Transportation from the airport to the China - EU Center for Agricultural Technology and hotel, a distance of about 20 km is available by taxi for about 10 US$ per car. Optional Post Symposium Tours Following the Symposium several tours will be arranged. The choices are: Route 1. one day tour, Beijing- Great Wall -Ming Tombs- Beijing. The cost is 30 US$ per person. Route 2. ten day tour,Beijing -Xiuran-Chongqing - Three Gorges-Guilin-Guangzhou. The cost is 1698 US$ per person. Route 3. six day tour, Beijing - Huangshang - Shanghai - Hangzhou- Shanghai. The cost is 935 US$ per person. Route 4. three day tour, Beijing - Taishan - Beijing. The cost is 366 US$ per person. Route 5. five day tour, Beijing - Xiuran - Guilin - Guangzhou. The cost is 860 US$ per person. All the above quotations are including hotel accommodation, daily ABF, Chinese lunch and dinner, domestic air and train fare, sightseeing entrance tickets, all land transportation, English-speaking guide. Participants interested in the tours please declare your interesting in the registration form. A non- refundable deposit is required. For more information please see Appendix and registration form. Registration Please return the enclosed registration form, payment, and abstract to: ISDVC c/o National Engineering Research Center for Vegetables P.O.Box 2443 Beijing 100081 China E-mail: huxg@bepc2.ihep.ac.cn / huxg@public.bta.net.cn Payment must be sent by check or Money Order in US Dollar and received by July 1, 1996 to qualify for pre- registration rate. Please mail questions regarding the Symposium to the Secretariat. Hu Xiaoguang National Engineering Research Center for Vegetables P.O.Box 2443 Beijing 100081 China Tel. ( +86 10 ) 8413217 or 8414433 Ext. 3083 Fax. ( +86 10 ) 8426286 E-mail: huxg@bepc2.ihep.ac.cn / huxg@public.bta.net.cn ********************************************************* ****** Registration Form Third International Symposium on Diversification of Vegetable Crops (ISDVC) September 24 - 27, 1996 Pre-registration deadline: July 1, 1996 Name in full ( last name first ) ________________________________________ Mailing address__________________________________________________ _ __________________________________________________ Telephone _______________________ Fax.__________________________ ___________ US$ 200 (US$ 160 for member of the ISHS )pre-registration including reception, breaks, general tour, evening entertainment, conference materials and proceedings. ____________ US$ 250 late registration ( after July 1, 1996 ) ____________ US$ 50 spouse / guest fee ( including reception & general tour) Name(s): ________________________________ ____________ US$ 50 Hotel reservation deposit Date of arrival __________ will stay________ days. _________ US$ 30, Route 1, Beijing -Great Wall - Ming Tombs _________ Number of persons attending. _________ US$ 200, Route 2 deposit, Beijing - Xiuran - Three Canyon-Guilin - Guangzhou. _________ Number of persons attending _________ US$ 150, Route 3 deposit, Beijing - Huangshan-Shanghai-Hangzhou- Shanghai.________ Number of persons attending. _________ US$ 100, Route 4 deposit, Beijing - Taishang - Beijing. ________ Number of persons attending. _________ US$ 150, Route 5 deposit, Beijing- Xiuran- Guilin-Guangzhou. _________ Number of persons attending. _________ Total enclosed Check or Money Order will be accepted, payable to: The Regents, National Engineering Research Center for Vegetables ( US dollars only ) Refund policy: Full refund will be issued upon cancellation prior to August 10, 1996. After that date only 50% of the registration fee will be refundable and the deposit for post symposium tours is non-refundable. Please mail this form with payment to: ISDVC c/o National Engineering Research Center for Vegetables P.O.Box 2443 Beijing 100081 China Pre - registration deadline - July 1, 1996 ********************************************************* *************** Appendix: OPTIONAL POST SYMPOSIUM TOURS Route 1: Beijing - Great Wall - Ming Tomb - Beijing * Price: US$ 30 Sep. 28 One day tour. Route 2: Beijing-Xi'an-Chongqing-Three George-Guilin- Guangzhou-Exit * Price: US$ 1698.00 * Single room supplement: US$ 765.00 * Itinerary: Sep.28 Morning flight from Beijing to Xi'an Afternoon: Sightseeing to Old City Wall, Banpo Museum Stay in Dynasty Hotel(4 Star) Sep.29 Visit Terra-cotta Warriors, Huaqing Hot Spring Sep.30 Xi'an to Chongqing by air, visit E'ling Park Stay in Yangxijiang Holiday inn(4 Star) Oct.1-Oct 3 Yangzi River Cruise (Three George) by boat Oct.4 Arrival in Wuhan Stay in Changjiang Hotel(4 Star) Oct.5 Wuhan to Guilin by morning flight Afternoon: Sightseeing to Reed Cave Stay in Guilin Holiday Inn (4 Star) Oct.6 Cruise on Li River Oct.7 Guilin to Guangzhou by air Visit Chen's Temple, Dr. Sun Yat-Sen's Memorial Hall Stay in Jiangnan Hotel(4 Star) Oct.8 Exit Route 3: Beijing-Huangshan-Shanghai-Hangzhou-Shanghai- Exit * Price: US$ 935.00 * Single room supplement: US$ 225.00 * Itinerary: Sep.28 Beijing fly to Huangshan(Yellow Mountain) Climb up the mountain by cable-car Stay in Western-Sea Hotel(4 Star) Sep.29 Sightseeing in Huangshsn Sep.30 Huangshan to Shanghai by air Stay in Jianguo Hotel(4 Star) Oct. 1 Shanghai to Hangzhou by train Stay in Huagang Hotel(3 Star) Oct. 2 Visit the Western Lake, Lingyin Temple Oct. 3 Hangzhou to Shanghai by train Visit Yu Garden, Jade Buddha Temple Stay in Jianguo Hotel Oct.4 Exit Route 4: Beijing-Taishan-Beijing-Exit * Price: US$ 366.00 * Single room supplement: US$ 80.00 * Itinerary: Sep.28 Beijing to Tai'an by train Stay in Taishan Hotel Sep.29 Sightseeing to Mount Tai Sep.30 Back to Beijing Stay in Tiantan Hotel Oct. 1 Exit Route 5: Beijing-Xi'an-Guilin-Guangzhou-Exit * Price: US$ 860.00 * Single room supplement: US$ 138.00 * Itinerary: Sep.28 Beijing to Xi'an by morning flight Afternoon: City Wall, Banpo Museum Stay in Dynasty Hotel(4 Star) Sep.29 Visit Terra-cotta Warriors, Huaqing Hot Spring Sep.30 Xi'an to Guilin Holiday Inn Oct. 1 Cruise on Guilin to Guangzhou Visit Chen's Temple, Dr. Sun Yat-Senurs Memorial Hall Stay in Jiangnan Hotel(4 Star) Oct. 3 Exit All of the above quotations are including hotel accommodation, daily ABF, Chinese lunch and dinner, domestic air and train fare, sightseeing entrance tickets, all land transportation, English-speaking guide. ISDVC China - EU Center for Agricultural Technology Beijing, China 24 - 27 September, 1996 ISHS From owner-rapd@net.bio.net Wed Mar 06 22:00:00 1996 Path: biosci!ns1.faseb.org!lamarck.sura.net!newsfeed.internetmci.com!uwm.edu!cs.utexas.edu!news.tamu.edu!news From: lazo@tamu.edu (Gerard R. Lazo) Newsgroups: bionet.molbio.rapd Subject: Re: rapds in cotton Date: 6 Mar 1996 15:32:40 GMT Organization: USDA-ARS Southern Crops Research Laboratory Lines: 32 Distribution: world Message-ID: <4hkb6o$kfi@news.tamu.edu> References: <313C6B19.1F9C@comp.uark.edu> Reply-To: lazo@tamu.edu NNTP-Posting-Host: algodon.tamu.edu You can check out a cotton-related RAPD paper at: http://algodon.tamu.edu/homepage/lazo/docs/COT.html There is also some RAPD information listed in the Cotton Genome Database (CottonDB) located at: http://probe.nalusda.gov/ If you have any information that you can contribute to CottonDB, please use the forms located at: http://algodon.tamu.edu/ > From jamiles@COMP.UARK.EDU (jay miles) > Newsgroups: bionet.molbio.rapd > Subject: rapds in cotton > Date: Tue Mar 05 09:51:04 CST 1996 > Organization: U of AR > > I have a friend interested in doing rapd research with cotton. If anyone > knows of any useful protocols or primers, I'm sure he would be interested > in hearing from you. Contact Altaf Khan at makhan@comp.uark.edu ! > > Thanks -- Gerard R Lazo .-.-. USDA-ARS Southern Crops Research Laboratory -. .-. .-. .-. .-. ).). .-. .-. .-. .-. .-. .-. .-. .-. .-. ||X|||\ /|||X|||\(.( .) .)\ /|||X|||\ /|||X|||\ /|||X|||\ /|||X|||\ /||| |/ \|||X|||/ \|||X(_( ._)||X|||/ \|||X|||/ \|||X|||/ \|||X|||/ \|||X|||/ ' `-' `-' `-' `-//\)`-' `-' `-' `-' `-' `-' `-' `-' `-' `-' lazo@tamu.edu (( 2765 F&B Rd, College Station, TX 77845-9592 From owner-rapd@net.bio.net Wed Mar 06 22:00:00 1996 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!elroy.jpl.nasa.gov!swrinde!gatech!taco.cc.ncsu.edu!news From: Susan Jane Hogarth Newsgroups: bionet.molbio.rapd Subject: Re: Thermostability of Taq polymerase Date: 7 Mar 1996 16:13:14 GMT Organization: North Carolina State University Lines: 31 Message-ID: <4hn1uq$ni1@taco.cc.ncsu.edu> References: <552A2914F5A@plantpath.pp.ksu.edu> NNTP-Posting-Host: sparc5c-2203gardnr.ppath.ncsu.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.12 (X11; I; SunOS 5.4 sun4m) X-URL: news:552A2914F5A@plantpath.pp.ksu.edu >> > I don't know the maximum temperature, but most experimenters use 94 - >> > 95 C as the maximum. Maybe this has already been addressed, but if Taq polymerase is so stable at high temps, why are people so paranoid about it getting out of the freezer for more than 2.5 seconds? Just a question that's always bugged me.... -- Susan Jane Hogarth "Luck is the residue of design." -- Freddy the Fish "Personally, I'm always ready to learn, although I do not always like being taught." -- Winston Churchill http://www4.ncsu.edu/~sjhogart/public/home.html . .-~\ / `-'\.' `- : | / `._ | | .-. { \ | `-' `. . \ | / ~-.`. \| .-~_ `.\-.\ .-~ \ `-'/~~ -.~ / .-~/|`-._ /~~-.~ -- ~ / | \ ~- . _\ From owner-rapd@net.bio.net Wed Mar 06 22:00:00 1996 Path: biosci!daresbury!not-for-mail From: S P Harrison Newsgroups: bionet.molbio.rapd Subject: Wolf Date: 7 Mar 1996 14:01:54 -0000 Lines: 4 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <4hmq8i$obm@mserv1.dl.ac.uk> Original-To: rapd@dl.ac.uk I would be grateful if anybody could lwet or even let me have the telephone no. of a company called Wolf who supply electrophoresis eqt. etc. Thank you. S.P.Harrison From owner-rapd@net.bio.net Thu Mar 07 22:00:00 1996 Path: biosci!CS.Arizona.EDU!noao!ennfs.eas.asu.edu!gatech!newsfeed.internetmci.com!mr.net!news.mr.net!cruise.biol.stthomas.edu!jlcruise From: JLCruise Newsgroups: bionet.molbio.rapd Subject: Analyzing RAPD gels Date: 8 Mar 1996 21:49:22 GMT Organization: University of St. Thomas Lines: 11 Distribution: world Message-ID: <4hqa12$829@news.mr.net> NNTP-Posting-Host: cruise.biol.stthomas.edu Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1 Content-Transfer-Encoding: 8bit X-Newsreader: Nuntius 2.0.4_PPC X-XXMessage-ID: X-XXDate: Fri, 8 Mar 1996 20:51:06 GMT Is there free/share/inexpensive software available to analyze banding patterns on RAPD gels? We are interested in decision-making about paternity, etc., in samples from birds. Any info would be appreciated; please respond by email to: mjdejong@stthomas.edu Thank you. Mike DeJong University of St. Thomas From owner-rapd@net.bio.net Thu Mar 07 22:00:00 1996 Path: biosci!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!news.uoknor.edu!news.nodak.edu!plains!rai From: rai@plains.nodak.edu (Satish Rai) Newsgroups: bionet.molbio.rapd Subject: Taq polymerase Date: 8 Mar 1996 16:10:49 GMT Organization: North Dakota Higher Education Computing Network (NDHECN) Lines: 1 Message-ID: <4hpm69$lk9@daily-planet.nodak.edu> NNTP-Posting-Host: plains.nodak.edu Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit X-Newsreader: TIN [version 1.2 PL2] From owner-rapd@net.bio.net Fri Mar 08 22:00:00 1996 Path: biosci!daresbury!yama.mcc.ac.uk!sunsite.doc.ic.ac.uk!dundee.ac.uk!power.medschool.dundee.ac.uk!pmkeig From: pmkeig@ninewells.dundee.ac.uk (Paula Keig) Newsgroups: bionet.molbio.rapd Subject: Life expectancy of DNA Date: Sat, 9 Mar 1996 17:03:48 GMT Organization: University of Dundee Lines: 8 Message-ID: NNTP-Posting-Host: power.medschool.dundee.ac.uk X-Newsreader: Trumpet for Windows [Version 1.0 Rev A] I'm an honours student and am having just afew problems with my PCR rapids. can anyone help me, PLEASE!!! I am using DNA that was extracted in oct/nov 95.(stored in the fridge) up until about 4 weeks ago everything was ok, then all of a sudden I was getting no results. I've tried everything, different concentrations of DNA, different primers. I have one week to go on my project, so would like some help soon. Ant suggestions would be great. From owner-rapd@net.bio.net Fri Mar 08 22:00:00 1996 Path: biosci!LUXOR.LATROBE.EDU.AU!gendjc From: gendjc@LUXOR.LATROBE.EDU.AU (CLANCY David) Newsgroups: bionet.molbio.rapd Subject: PCR product detecting-quick and cheap please Date: 9 Mar 1996 15:53:44 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 15 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Morning, could anyone help me with a quick and cheap method of blotting PCR products (just to detect presence or absence of product) using coloured probes rather than radioactive. Emphasis on quick and cheap. I'm looking for an alternative to running minigels on about a thousand samples. Reply to my address and I'll broadcast the responses in one go if they sound like they'd be useful to more people than just me. Thanks again, rapdnet, and keep up the good work. The citizens can sleep safe in the knowledge etc etc etc etc. Dave Clancy Genetics Dept. La Trobe Univ. Bundoora 3083 From owner-rapd@net.bio.net Fri Mar 08 22:00:00 1996 Path: biosci!agate!howland.reston.ans.net!swrinde!gatech!taco.cc.ncsu.edu!faeppc2.cvm.ncsu.edu!user From: david_ley@ncsu.edu (David Ley) Newsgroups: bionet.molbio.rapd Subject: Gel Analysis - DNA ProScan Date: Sat, 09 Mar 1996 09:44:41 -0500 Organization: NCSU-CVM Lines: 5 Message-ID: NNTP-Posting-Host: faeppc2.cvm.ncsu.edu Greetings, Does anyone have experience with a gel (esp. RAPD) analysis program called DNA ProScan? I would be interested in your impressions of this software and info. on how to contact the company. Thanks in advance for your help, dl. From owner-rapd@net.bio.net Sat Mar 09 22:00:00 1996 Path: biosci!CATI.CSUFRESNO.EDU!aelashr From: aelashr@CATI.CSUFRESNO.EDU (Abd elnasser Elashry) Newsgroups: bionet.molbio.rapd Subject: asking about softwares Date: 10 Mar 1996 06:38:14 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 16 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603101437.GAA04118@CATI.CSUFresno.EDU> NNTP-Posting-Host: net.bio.net Hello everybody, I received very important email to me which have information about the soft ware and how anyone can get it free from the internet. this was about the soft ware which is used to analyze agarose gels. In fact i tried to save it but I can't and unfortunatelly lost it. so, if any one could send it to me again I'll be very appriciate. Thanks alot for your cooperation. Yours abd elnasser ***************************************************** Abd El-nasser Mohamed Abd El-Moneim Elashry Agricultural Research Center (ARC). Agricultural Genetic Engineering Research Institute (AGERI). 9 Gamaa St., Giza 12619 Egypt ***************************************************** From owner-rapd@net.bio.net Sat Mar 09 22:00:00 1996 Path: biosci!news.alaska.edu!news From: "James W. Fetzner Jr." Newsgroups: bionet.molbio.rapd Subject: RE: Life Expectancy of DNA Date: 10 Mar 1996 02:06:35 GMT Organization: University of Alaska Computer Network Lines: 52 Message-ID: <4htdfb$59f@news.alaska.edu> NNTP-Posting-Host: 146.63.246.34 Paula Keig wrote: >I'm an honours student and am having just afew problems with my PCR rapids. >can anyone help me, PLEASE!!! >I am using DNA that was extracted in oct/nov 95.(stored in the fridge) >up until about 4 weeks ago everything was ok, then all of a sudden I was >getting no results. I've tried everything, different concentrations of DNA, >different primers. I have one week to go on my project, so would like some >help soon. >Ant suggestions would be great. Dear Paula, Unfortunately, it sounds like your DNA may have degraded while in storage. This would especially be suspect if you rehydrated your samples in water and not some sort of buffering agent and if you amplified your samples before and they don't work now. I have found that when DNA is rehydrated in water and stored at 4C, the life expectancy of the DNA sample is greatly diminished (on the order of a few weeks to a few months) and PCR of very large or moderately large (>1000 bp) fragments falls apart pretty quickly. If this is the procedure you followed, you may have to re-extract your samples. Perhaps the quickest way of checking this would be to run some of your samples (stock DNA) on a agarose gel and checking the size of your DNA against known size standards (usually lambda DNA). If you have no high molecular weight DNA, re-extracting is definitely the way to go (both in terms of time and expense when compared to continued PCR troubleshooting). If degridation is the problem, it won't matter how much template DNA (from your current stocks) or primer you add to the PCR reaction, the samples just won't amplify well, if at all, especially if your RAPD products are large! The best way I have found to avoid this problem is to rehydrate your DNA samples in either TE (10 mM Tris, 0.1mM EDTA; pH 8.0) or just in 10mM Tris, pH 8.0. The EDTA in the TE buffer above does not seem to interfere with PCR, even at MgCl2 concentrations as low as 1.5 mM. I routinely store DNA at 4C in TE buffer and have yet to have a problem with degridation of my DNA samples (even those over a year and a half old), and easily amplify 2.5 kb fragments from these samples. Hope this has been of some help to you. Sincerely, James W. Fetzner Jr. Project Geneticist Alaska Dept. of Fish & Game Genetics Lab Anchorage, AK From owner-rapd@net.bio.net Sun Mar 10 22:00:00 1996 Path: biosci!CS.Arizona.EDU!noao!ennfs.eas.asu.edu!gatech!newsfeed.internetmci.com!uwm.edu!vixen.cso.uiuc.edu!usenet.ucs.indiana.edu!willow.bio.indiana.edu!user From: dewolf@bio.indiana.edu (Diana Wolf) Newsgroups: bionet.molbio.rapd Subject: Re: Life expectancy of DNA Date: 11 Mar 1996 00:54:53 GMT Organization: Indiana University Lines: 26 Message-ID: References: NNTP-Posting-Host: willow.bio.indiana.edu In article , pmkeig@ninewells.dundee.ac.uk (Paula Keig) wrote: > I'm an honours student and am having just afew problems with my PCR rapids. > can anyone help me, PLEASE!!! > I am using DNA that was extracted in oct/nov 95.(stored in the fridge) > up until about 4 weeks ago everything was ok, then all of a sudden I was > getting no results. I've tried everything, different concentrations of DNA, > different primers. I have one week to go on my project, so would like some > help soon. > Ant suggestions would be great. If your problem is DNA degredation, I would expect that the large bands would slowly dissapear, gradually leading to lighter and lighter bands. You wouldn't suddenly cease to get any reaction. I would first throw away all of my solutions, and make new ones in different, sterilized containers. The ones you are using may be contaminated. If this doesn't work, try some new taq. Maybe someone left it out of the freezer for too long. Diana -- Diana Wolf Biology Department Indiana University From owner-rapd@net.bio.net Sun Mar 10 22:00:00 1996 Path: biosci!ACD.TUSK.EDU!Prakash From: Prakash@ACD.TUSK.EDU (C. S. Prakash) Newsgroups: bionet.molbio.rapd Subject: US Residential Phone Numbers Now Available on Internet! Date: 10 Mar 1996 18:05:49 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 23 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net There is a new free new service where you can get phone number of virtually any listed telephone number along with the address in the U. S. through a Internet site. Point your web browser to http://www.switchboard.com/ The database contains 90 millions phone numbers of residences and companies, and also email addresses of registered users. It is an amazing and helpful database but sure to be nuisance to celebrities. I tried a few Nobel Laureates names and found a few of them listed! -+*%$%*+-+*%$%*+-+*%$%*+-+*%$%*+-+*%$%*+--+*%$%*+- C. S. Prakash Phone (334) 727 8023 Tuskegee University Fax (334) 727 8552 School of Agriculture Email: Prakash@Acd.Tusk.Edu Tuskegee, AL 36088. USA -+*%$%*+-+*%$%*+-+*%$%*+-+*%$%*+-+*%$%*+--+*%$%*+- From owner-rapd@net.bio.net Mon Mar 11 22:00:00 1996 Path: biosci!labatt.com!rob.stewart From: rob.stewart@labatt.com ("Stewart, Robert") Newsgroups: bionet.molbio.rapd Subject: FW: Life expectancy of DNA Date: 12 Mar 1996 13:37:17 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 30 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Greetings, We have experienced similar problems with yeast DNA. Generally we have to dilute the DNA for storage at 4 C, otherwise it degrades rapidly, presumably due to residual nuclease activity. In your case perhaps your DNA has been contaminated by enzymes/bacteria. I suggest that you precipitate the DNA with ethanol. This may help the RAPD pattern. Bare in mind that the RAPD pattern of the DNA may have changed due to partial breakdown and the previous results may not be recoverable. You might have to live with having less then 100% concordance with previous experiments. Cheers RS ---------- From: pmkeig@ninewells.dundee.ac.uk[SMTP:pmkeig@ninewells.dundee.ac.uk] Sent: Saturday, March 09, 1996 12:03PM To: rapd@net.bio.net Subject: Life expectancy of DNA I'm an honours student and am having just afew problems with my PCR rapids. can anyone help me, PLEASE!!! I am using DNA that was extracted in oct/nov 95.(stored in the fridge) up until about 4 weeks ago everything was ok, then all of a sudden I was getting no results. I've tried everything, different concentrations of DNA, different primers. I have one week to go on my project, so would like some help soon. Ant suggestions would be great. From owner-rapd@net.bio.net Mon Mar 11 22:00:00 1996 Path: biosci!YVAX.BYU.EDU!anderswr From: anderswr@YVAX.BYU.EDU (W. Ralph Andersen) Newsgroups: bionet.molbio.rapd Subject: (none) Date: 12 Mar 1996 14:05:22 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 18 Sender: daemon@net.bio.net Distribution: world Message-ID: <01I297AX20IUB12QGF@yvax.byu.edu> NNTP-Posting-Host: net.bio.net Visiting scientists at our lab, Paulo Ruas and Claudette Ruas are looking for a good used Perkin Elmer 48-well thermocycler----or a good used demonstrator of this size. We are looking for one that works well or one that can be repaired. I would greatly appreaciate any help you folks can give us. Thanks. Ralph Say, thanks for your replies on my quest for help on my power PC 9500. Many very useful suggestions. Thanks again. Dr. W. Ralph Andersen 297 WIDB Brigham Young University Provo, UT 84602 Ph. 801 378 2468 Fax 801 378 7499 From owner-rapd@net.bio.net Tue Mar 12 22:00:00 1996 Newsgroups: bionet.molbio.rapd Path: biosci!CS.Arizona.EDU!noao!ennfs.eas.asu.edu!gatech!newsfeed.internetmci.com!tank.news.pipex.net!pipex!peer-news.britain.eu.net!strath-cs!info!cmmeghji From: cmmeghji@exeter.ac.uk (C.M.Meghji) Subject: Re: Life expectancy of DNA Message-ID: Organization: University of Exeter, UK References: Date: Wed, 13 Mar 1996 12:36:28 GMT Lines: 13 - sorry to hear that you are suffering from no products in your RAPD (NOT rapid, by the way there is nothing particularly fast about it!). This phenomenon is the bane of the the RAPD msappers life and it has just descended on you, but don't feel picked on it happens to EVERYBODY. The shortest answer is - Discard Everything and start again from scratch. Your DNA is probably not the problem, get new: water, taq,dNTPs,Mg++ oil etc. Make up your dNTP,s and whatever in NEW water, and make new DNA diluitions from your base stock, if you have it. Replace as much as you can, don't try to eliminate individual elements as the cause of the problem at this stage. Good luck, and welcome to the club Carol Meghji From owner-rapd@net.bio.net Tue Mar 12 22:00:00 1996 Path: biosci!IRRI.CGNET.COM!NHUANG From: NHUANG@IRRI.CGNET.COM Newsgroups: bionet.molbio.rapd Subject: DNA sequence analysis software Date: 13 Mar 1996 15:09:33 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 10 Sender: daemon@net.bio.net Distribution: world Message-ID: <01I2BJ5KUYTU8WX3GU@IRRI.CGNET.COM> NNTP-Posting-Host: net.bio.net We have partially sequenced several cloned RAPD fragments and wish to analyze the sequence to design STS primers. Anybody knows where I can get a DNA sequence analysis sofeware. Thanks in advance. Ning Haung E-mail Nhuang@CGNET.COM FAX 1-415-8336621 Tel 1-415-8336620 From owner-rapd@net.bio.net Wed Mar 13 22:00:00 1996 Path: biosci!CATI.CSUFRESNO.EDU!aelashr From: aelashr@CATI.CSUFRESNO.EDU (Abd elnasser Elashry) Newsgroups: bionet.molbio.rapd Subject: needing to more explanations Date: 14 Mar 1996 10:12:34 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 13 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603141811.KAA08873@CATI.CSUFresno.EDU> NNTP-Posting-Host: net.bio.net Dear netters: I am a master student, and my project is about Molecular characterization of Meloidogyne. I heard from more than one that the RAPD PCR is not reliable to detect the relatedness of organisms, because of many reasons such as:"the Results are easy to be changed by changing any conditions, and that may be happen many times, that is one of the reasons". I don't know is that right or not!, So that, if anybody there can help me to determine the usefulness of this method to my work comparing with other methods, it will be very useful to me. I am waiting for your opinions. Thanks for your kindness and cooperation. Yours Abd Elnasser From owner-rapd@net.bio.net Wed Mar 13 22:00:00 1996 Path: biosci!bcm.tmc.edu!pendragon!news.msfc.nasa.gov!elroy.jpl.nasa.gov!swrinde!howland.reston.ans.net!newsfeed.internetmci.com!info.ucla.edu!library.ucla.edu!csulb.edu!csus.edu!news.ucdavis.edu!ALPHA.UCDAVIS.EDU!BAGLEY From: bagley@ALPHA.UCDAVIS.EDU Newsgroups: bionet.molbio.rapd Subject: Stoffel Fragment and RAPD band size Date: 14 Mar 1996 01:51:35 GMT Organization: Animal Science Dept., UC Davis, Davis, CA Lines: 8 Message-ID: <4i7u37$pfi@mark.ucdavis.edu> Reply-To: bagley@ALPHA.UCDAVIS.EDU NNTP-Posting-Host: alpha.ucdavis.edu Does anyone know of a reference that specifically states that the use of Stoffel fragment Taq polymerase results in RAPD profiles with smaller average band size than when normal Taq is used? I am aware of one (anonymous) Perkin-Elmer brochure but I am reluctant to cite it. Thanks Mark bagley@ansci.ucdavis.edu From owner-rapd@net.bio.net Wed Mar 13 22:00:00 1996 Path: biosci!ACD.TUSK.EDU!Prakash From: Prakash@ACD.TUSK.EDU (C. S. Prakash) Newsgroups: bionet.molbio.rapd Subject: Re: Stoffel Fragment and RAPD band size Date: 14 Mar 1996 08:30:35 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 33 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net >Does anyone know of a reference that specifically states that the use of >Stoffel fragment Taq polymerase results in RAPD profiles with smaller >average band size than when normal Taq is used? I am aware of one >(anonymous) Perkin-Elmer brochure but I am reluctant to cite it. Mark: Try these two references which mention the use of Stoffel v/s untruncated polymerases in the RAPD analysis Aldrich, J. and C. Cullis. 1993. RAPD analysis in flax: Optimization of yield and reproducibility using KlenTaq 1 DNA polymerase, Chelex 100 and gel purification of genomic DNA. Plant Mol. Bio. Reporter 11:128-141. Sobral, B.W.S. and R. Honeycutt. 1993. High output genetic mapping of polyploids using PCR-generated markers. Theor. Appl. Genet. 86:105-112. --------------------------------------------- C. S. Prakash Prakash@Acd.Tusk.Edu Tuskegee University Phone (334) 727 8023 School of Agriculture Fax (334) 727 8552 Tuskegee, AL 36088. USA ---------------------------------------------- From owner-rapd@net.bio.net Wed Mar 13 22:00:00 1996 Path: biosci!rutgers!gatech!newsfeed.internetmci.com!howland.reston.ans.net!nntp.coast.net!fu-berlin.de!zrz.TU-Berlin.DE!news.dfn.de!news.uni-jena.de!news From: Johannes Woestemeyer Newsgroups: bionet.molbio.rapd Subject: Genetic Meeting Jena Date: 14 Mar 1996 16:08:12 GMT Organization: FSU Jena: Inst. Gen. Microbiol. / Microbe Genetics Lines: 29 Message-ID: <4i9g9c$dga@fsuj19.rz.uni-jena.de> NNTP-Posting-Host: pc1.mikrogen.uni-jena.de Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.2N (Windows; I; 16bit) Dear colleagues, from September 23. to 26. we will have this year's meeting of the German Genetic Society in Jena. The programme covers many aspects of modern genetics, especially * Control of gene expression * Human genetics * Plants and plant diseases * Fungal systems * Developmental genetics You are cordially invited to take an active part in this meeting. The conference language will be english. You may contribute posters or suggestions for talks. Just drop me a note, and I will send you more information. Joh. Woestemeyer Address: FSU Jena Inst. General Microbiology/Microbe Genetics Neugasse 24 D-07743 Jena / Germany Tel: +49 (0)3641 630310 Fax: *49 (0)3641 630321 Email: b5wojo@rz.uni-jena.de From owner-rapd@net.bio.net Wed Mar 13 22:00:00 1996 Path: biosci!rutgers!uwm.edu!newsfeed.internetmci.com!howland.reston.ans.net!surfnet.nl!swsbe6.switch.ch!swidir.switch.ch!in2p3.fr!univ-lyon1.fr!jussieu.fr!saphir.jouy.inra.fr!usenet From: "Patrick.Tailliez" Newsgroups: bionet.molbio.rapd Subject: Re: Analyzing RAPD gels Date: 14 Mar 1996 13:24:39 GMT Organization: INRA Lines: 14 Message-ID: <4i96mn$oq6@saphir.jouy.inra.fr> References: <4hqa12$829@news.mr.net> NNTP-Posting-Host: saphir.jouy.inra.fr Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22 (Windows; I; 16bit) To: jlcruise@stthomas.edu Dear Mike, There are softwares available to analyse banding patterns on RAPD gels. One I actually use for that, was developped for SDS-PAGE patterns comparisons.It's GelCompar software distributed by Applied Maths, Kortrijk, Belgium, Fax +32 56 402 145. But it's not free ! This soft is interesting in RAPD patterns analysis and comparison because it takes into account the position of each band and its intensity. It is also able to make corrections of the gels. One reference : Analysis of electrophoretic whole-organism protein fingerprints. Pot et al, 1994 in Chemical Methods in Prokaryotic systematics, edited by Goodfellow and O'Donnell. Good luck ! Patrick TAILLIEZ Curator of the CNRZ Collection of Lactic Acid Bacteria From owner-rapd@net.bio.net Thu Mar 14 22:00:00 1996 Path: biosci!rutgers!gatech!newsfeed.internetmci.com!news.flinet.com!news1.inlink.com!news.starnet.net!wupost!news.utdallas.edu!news.tamu.edu!news From: lazo@tamu.edu (Gerard R. Lazo) Newsgroups: bionet.molbio.rapd Subject: Re: Stoffel Fragment and RAPD band size Date: 14 Mar 1996 16:45:16 GMT Organization: USDA-ARS Southern Crops Research Laboratory Lines: 37 Message-ID: <4i9ies$7qj@news.tamu.edu> References: <4i7u37$pfi@mark.ucdavis.edu> Reply-To: lazo@tamu.edu NNTP-Posting-Host: algodon.tamu.edu I compared the Stoffel fragment and holoenzyme while doing RAPDs on cotton. I have a picture (scan-impaired) of it available over the web at: http://algodon.tamu.edu/homepage/lazo/docs/COT.html The citation would probably be as such: Lazo, G.R., Park, Y.-H., and Kohel, R.J. 1994. Identification of RAPD markers linked to fiber strength in Gossypium hirsutum and G. barbadense interspecific crosses. Pages 71-77 in: Biochemistry of Cotton Workshop, Cotton Incorporated, September 28-30, 1994, Galveston, TX. 126 pp. -- Gerard R Lazo .-.-. USDA-ARS Southern Crops Research Laboratory -. .-. .-. .-. .-. ).). .-. .-. .-. .-. .-. .-. .-. .-. .-. ||X|||\ /|||X|||\(.( .) .)\ /|||X|||\ /|||X|||\ /|||X|||\ /|||X|||\ /||| |/ \|||X|||/ \|||X(_( ._)||X|||/ \|||X|||/ \|||X|||/ \|||X|||/ \|||X|||/ ' `-' `-' `-' `-//\)`-' `-' `-' `-' `-' `-' `-' `-' `-' `-' lazo@tamu.edu (( 2765 F&B Rd, College Station, TX 77845-9592 > From bagley@ALPHA.UCDAVIS.EDU > Newsgroups: bionet.molbio.rapd > Subject: Stoffel Fragment and RAPD band size > Date: Wed Mar 13 19:51:35 CST 1996 > Organization: Animal Science Dept., UC Davis, Davis, CA > > Does anyone know of a reference that specifically states that the use of > Stoffel fragment Taq polymerase results in RAPD profiles with smaller > average band size than when normal Taq is used? I am aware of one > (anonymous) Perkin-Elmer brochure but I am reluctant to cite it. > Thanks > Mark > > bagley@ansci.ucdavis.edu From owner-rapd@net.bio.net Thu Mar 14 22:00:00 1996 Path: biosci!labatt.com!rob.stewart From: rob.stewart@labatt.com ("Stewart, Robert") Newsgroups: bionet.molbio.rapd Subject: RE: needing to more explanations Date: 15 Mar 1996 12:22:27 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 41 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Greetings, It is correct that the patterns generated by RAPD PCR cannot be used for phylogenetic results. This is due to random and relatively non-specific nature of amplification. Strains of the same organism may have profoundly different patterns. That is, a single sequence change may have a large effect on the RAPD outcome. If one base pair change results in the addition of a new amplicon this may not only add a new band but it could affect the amplification of another amplicon resulting in its loss. Also, changes in the reaction efficiency due to differences in enzyme activity ( very important), DNA concentrations, or the presence of inhibitory substances can change which fragments will be amplified. Other methods such as ribosomal RNA amplifications with subsequent restriction enzyme analyses or direct sequencing are much more accurate measures of phylogeny. I hope this is helpful. Tallyhoe RS ---------- From: aelashr@cati.csufresno.edu[SMTP:aelashr@cati.csufresno.edu] Sent: Thursday, March 14, 1996 1:12PM To: rapd@net.bio.net Subject: needing to more explanations Dear netters: I am a master student, and my project is about Molecular characterization of Meloidogyne. I heard from more than one that the RAPD PCR is not reliable to detect the relatedness of organisms, because of many reasons such as:"the Results are easy to be changed by changing any conditions, and that may be happen many times, that is one of the reasons". I don't know is that right or not!, So that, if anybody there can help me to determine the usefulness of this method to my work comparing with other methods, it will be very useful to me. I am waiting for your opinions. Thanks for your kindness and cooperation. Yours Abd Elnasser From owner-rapd@net.bio.net Thu Mar 14 22:00:00 1996 Path: biosci!LIFSCI.SDSU.EDU!MCCLELLAND From: MCCLELLAND@LIFSCI.SDSU.EDU Newsgroups: bionet.molbio.rapd Subject: reproducibility and phylogenetics Date: 15 Mar 1996 15:29:02 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 12 Sender: daemon@net.bio.net Distribution: world Message-ID: <960315152651.2020154f@LIFSCI.SDSU.EDU> NNTP-Posting-Host: net.bio.net Robert Stewart makes two good points: One is that the loss or gain of a band will affect the other bands. However, we have pointed out that the more complex the pattern, the less effect the loss or gain of a band is. All our papers use fingerprints with in excess of 20 bands per primer. The "context effect" is not a problem in such a case. As to reproducibility it is simple to use two different DNA concetrations, say 50 ng and 25 ng from each individual to ensure that the DNAs are of similar quality and quantitiy. Saddly, neither of these rules are followed, with the resulting opinion spread that the method is no good. It certainly is no good if you do not do it right. Michael McClelland From owner-rapd@net.bio.net Thu Mar 14 22:00:00 1996 Path: biosci!BIOTECH.UFL.EDU!billm From: billm@BIOTECH.UFL.EDU (Bill McKendree) Newsgroups: bionet.molbio.rapd Subject: FloriGen newsgroup keyword search Date: 15 Mar 1996 06:54:49 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 48 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net ANNOUNCING: The Florida Genetics Corp. website (http://www.biotech.ufl.edu/bdi/florigen) has just installed a prototype newsgroup keyword search feature. This database includes messages posted since November 95 to the following newsgroups: Methods, etc.: methods-and-reagents mol-evol rapd grasses molreps Journals: bio-jrnl Announcements: bionews The messages we add to this database are (mostly) responses to initial postings, hopefully to cut down on the number of unnecessary hits for any keyword. The database is updated weekly, and should be useful for locating protocols, product sources or complaints, and just general info about your topic of interest. Each file that results from a hit will contain all messages posted to that newsgroup (and perhaps others) for that day, so we recommend downloading the larger files. Please send comments and suggestions for improvement to: florigen@biotech.ufl.org Happy Hunting!! _______________________ Bill McKendree, PhD Florida Genetics Corp. 12085 Research Drive. Alachua, FL 32615 (904) 462-0895 phone (904) 462-0875 fax _______________________ From owner-rapd@net.bio.net Fri Mar 15 22:00:00 1996 Path: biosci!ns1.faseb.org!lamarck.sura.net!csc.mc.edu!usenet From: "Robert G. Hamilton" Newsgroups: bionet.molbio.rapd Subject: Re: reproducibility and phylogenetics Date: 16 Mar 1996 17:15:45 GMT Organization: Mississippi College Lines: 17 Message-ID: <4iet01$d49@csc.mc.edu> References: <960315152651.2020154f@LIFSCI.SDSU.EDU> NNTP-Posting-Host: mobo.mc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Windows; I; 16bit) Michael McClelland makes a good point about reproducability. RAPDs seems to suffer more from imagined problems than real problems. In many cases the real problem is a lack of rigor. While you cannot assume that a specific pair of co-migrating bands are homologus, it seems that in general, you can say that co-migrating bands are homologous, and include a large number of bands. We run each sample at least three times. We complete an entire analysis and then use our conclusions as hypotheses that are tested with another round of sampling. (I use RAPDs for assessing genetic diversity within populations) Any method has limitations. The key is to work well within those limitations. If I were using RAPDs for phylogenetic analyses, I would want to sequence the bands that were critical to my analysis to test homology. From owner-rapd@net.bio.net Fri Mar 15 22:00:00 1996 Path: biosci!rutgers!gatech!newsfeed.internetmci.com!battery.awod.com!harbour.awod.com!usenet From: "Arthur Q. Villordon" Newsgroups: bionet.molbio.rapd Subject: Re: Stoffel Fragment and RAPD band size Date: 16 Mar 1996 09:27:22 GMT Lines: 8 Message-ID: <4ie1hq$i4m@harbour.awod.com> References: <4i7u37$pfi@mark.ucdavis.edu> NNTP-Posting-Host: chs0063.awod.com Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22KIT (Windows; U; 16bit) To: bagley@ALPHA.UCDAVIS.EDU Try and look up Sobral and Honeycutt. 1993. TAG 86:105-112. Hope this helps (and my citation is correct). Arthur ******************************* From owner-rapd@net.bio.net Sat Mar 16 22:00:00 1996 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.rapd Subject: IMPORTANT - BIOSCI Fundraising Update! Date: 17 Mar 1996 02:00:31 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 149 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603171000.CAA06575@net.bio.net> NNTP-Posting-Host: net.bio.net I'm interrupting the usual monthly posting of the BIOSCI miniFAQ to bring you up to date on BIOSCI fundraising progress, a topic of concern to your future use of this resource. Thank you in advance for taking the time to read this message carefully. Last year we announced that BIOSCI was going to adopt the U.S. Public Broadcasting System model to fund its operations after our DOE/NSF grant runs out later this year. Unlike PBS, we are not soliciting contributions from users; we are only selling ads on our Web pages solely to cover our operating costs. Our goal is to seek sponsorships until we build up an operating reserve of about $100,000 and then cease further promotions until we need to build the reserve back up. (The accountants among our readership will be familiar with the problem of deferred revenue which we can not safely utilize until ads have been displayed for a period of time.) We have three sponsors to date with a couple more pending. The process is time-consuming, however, and we need your help as explained further below. Our operating costs consist of our network connection, phone lines, hardware maintenance (we hope to have new and faster hardware soon!), plus 0.7 FTE of salaries covering UNIX systems admin, technical support, quality assurance, i.e., testing, of our system, and administrative costs (such as the time it takes to actually find/write/call potential sponsors and raise money!). Although the BIOSCI staff does get compensated for a portion of the work that they do, this project has always received a lot of free after-hours and "vacation" time labor, so we hope that no one will begrudge the time that we do charge to the project to serve you. All of the three part-time staff members, Dave Mack, Julie Lawrence, and myself, have full time day jobs and families in addition to working hard to keep this service running for all of you. Julie and Dave Mack are subcontractors for BIOSCI; my time that is charged to the project defrays a portion of my regular salary instead of adding to my income. Besides having to relocate the project, we were very busy this last year building new infrastructure such as our WWW hypermail interface to the system. This was released last December along with scores of WAIS indices for the newsgroups. Virtually everything is complete, although we do continue to find and fix bugs (many through your helpful feedback!). We are still having some problems with our WAIS indexing. The archives continue to grow rapidly. We are running over 100 indexes now versus three previously and any systems crashes cause greater havoc with the indexing than before! We are still working to fix this as fast as our resources permit and appreciate your patience, but we have been able to automate a lot of the infrastructure to reduce labor as compared to past requirements. We have also implemented new software to make moderation of BIOSCI/bionet newsgroups much easier and combat the growing problem of Internet junk mail and USENET "spamming." About 20% of our groups are now moderated, many of them by the BIOSCI staff! This, for example, made a major difference last year in the quality of content in our EMPLOYMENT/bionet.jobs.offered newsgroup which many commercial concerns and recruiting firms are using **without charge** to recruit candidates for positions in the biological sciences. We are also now in a position to have sponsors for individual newsgroups as you will have noticed if you have visited http://www.bio.net/ and clicked on "Access the BIOSCI/bionet newsgroups" recently. So, how can you help?? ---------------------- As noted above it can take a lot of time to contact potential sponsors if I have to do it all myself. Our request is quite simple. You can do two important things which will take very little time for you individually. First, please use our WWW system at http://www.bio.net/ to access the archives. You can now post or reply to messages via your Web browser. Your usage helps attract sponsors. If you contact any of our sponsors, please be sure to thank them for supporting BIOSCI. It is critical for them to get this feedback if they are to continue their sponsorship for the long term. Second, if you work for a company or organization that provides products or services of interest to the biology community, please pass this message on to your marketing or marketing communications department or other appropriate group. Please ask them to help support BIOSCI by sponsoring our Web site and explain the uses and benefits of the system to the biology community. If they are interested, they can then contact us for further information at our tech support address, biosci-help@net.bio.net. Our hope is to quickly raise several large corporate/institutional sponsors on our heavily-used WWW locations (some stats appended below), and then end this sponsorship campaign so that our resources can continue to be used for service provision, not fundraising. Many of our specialty newsgroup WWW archives are still used by small communities of scientists (and they haven't been heavily promoted yet). While these may be valuable niche markets to some advertisers, it will generate more labor and overhead having to find these sponsors, fairly price the locations, and deal with lots of smaller sponsorships than fewer mid-to large sponsors. We are striving to keep our operation as lean and efficient as possible since we are not trying to make careers out of running BIOSCI. We are trying if at all possible to avoid the administrative overhead entailed with processing lots of small payments to reach our fundraising goals. I'd like to thank all of you for your help in advance. In helping us, you are also helping yourselves, not only in keeping this resource available for all of the both large and small research communities that we serve, but also by alleviating the need for us to go back and compete with researchers for tight grant dollars! We promised NSF when we were awarded the BIOSCI grant that we would carry out this mission to make the service self-supporting. With your help, we will succeed in continuing BIOSCI's work into its second decade. Thank you very much! Sincerely, Dave Kristofferson BIOSCI/bionet Manager biosci-help@net.bio.net A list of our prime WWW sponsorship locations follow. Statistics are for the four week period from 22 Jan. - 18 Feb. 1996 and usage continues to grow. ---------------------------------------------------------------------- The overall BIOSCI WWW pages are currently visited by users from close to 5000 unique computer hosts per week. Web servers only log the Internet computer/host name and frequently more than one individual can connect to us from a particular host. Main home page, http://www.bio.net, visited recently by about 2100 unique hosts per week Main Newsgroups archives page, http://www.bio.net/archives.html, visited recently by about 1200 Unique hosts per week BIO-JOURNALS archive page, http://www.bio.net/BIO-JOURNALS.html, visited recently by about 1000 unique hosts per week. EMPLOYMENT archive pages: http://www.bio.net:80/hypermail/EMPLOYMENT/ and monthly header pages, visited recently by about 600 unique hosts per week. Address database search page, http://www.bio.net/addrsearch.html, visited recently by about 450 unique hosts per week. Methods newsgroup archive pages, http://www.bio.net:80/hypermail/METHDS- REAGNTS/ and monthly header pages, visited recently by about 350 unique hosts per week. ---------------------------------------------------------------------- From owner-rapd@net.bio.net Sat Mar 16 22:00:00 1996 Path: biosci!ml.csiro.au!chris.bolch From: chris.bolch@ml.csiro.au (Chris Bolch) Newsgroups: bionet.molbio.rapd Subject: Re: reproducibility and phylogenetics Date: 17 Mar 1996 14:42:50 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 61 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603172239.JAA02006@aqueous.ml.csiro.au> NNTP-Posting-Host: net.bio.net Here Here Michael and Robert, Please note, capitals are for emphasis, not shouting!! Reproduceability is very good if care and attention is paid to the reaction conditions, reagents and DNA quality. I think there are now many examples where RAPD has confirmed/ mirrored results from more "reliable" methods, so it is clear to me that RAPD can be a reliable method for measuring genetic similarity/difference (which is what we do with all approaches) and then INFERRING phylogeny. One cannot come up with such concrete measures as "%sequence divergence" with RAPD but it is extreme to discount RAPD altogether. Robert Hamilton makes the very good point of working within the limitations of the method- a good example is within/ between popn analyses. When comparing divergent taxa I would expect that RAPD would become a little suspect. More problems are likely to be encountered with band homology etc. If the organisms are likely to be fairly divergent (e.g clearly different spp.) then why not choose a more "conventional" tool which will undoubtedly show enough variation if the former is a realistic hypothesis. IF it really bothers you then sequnece and make some STS's (SCARS, or whatever you want to call them). The problem with RAPD still lies in interpretation. Once there is a better understanding of the relationship between bands within and between strains it will be clearer which is the most appropriate approach for scoring and analysis. The keyword to use is CAUTION when interpreting RAPD analyses, and always try to obtain congruent data via a different approach if possible. Chris >Robert Stewart makes two good points: One is that the loss or gain of a >band will affect the other bands. However, we have pointed out that the >more complex the pattern, the less effect the loss or gain of a band is. >All our papers use fingerprints with in excess of 20 bands per primer. >The "context effect" is not a problem in such a case. As to reproducibility >it is simple to use two different DNA concetrations, say 50 ng and 25 ng >from each individual to ensure that the DNAs are of similar quality and >quantitiy. Saddly, neither of these rules are followed, with the resulting >opinion spread that the method is no good. It certainly is no good if you >do not do it right. > >Michael McClelland ________________________________________ Any views, statements of intent, or comment in this letter have no official status and do not constitute or imply any contract with myself or my employer. ________________________________________ Christopher J. S. Bolch, Algal Ecologist/Geneticist, CSIRO Division of Fisheries, GPO Box 1538, Hobart, Tasmania, Australia, 7001. PH. 002 325314 (Aus), 061-02-325314 (Internat.) FAX. 002 325000 (Aus), 061-02-325000 (Internat.) ________________________________________ From owner-rapd@net.bio.net Sun Mar 17 22:00:00 1996 Path: biosci!agate!howland.reston.ans.net!vixen.cso.uiuc.edu!uwm.edu!reuter.cse.ogi.edu!engr.orst.edu!newsfeed.orst.edu!news.orst.edu!ava.bcc.orst.edu!pomperk From: Kirk Pomper Newsgroups: bionet.molbio.rapd Subject: Re: Life expectancy of DNA Date: Mon, 18 Mar 1996 11:27:36 -0800 Organization: University Computing Services - Oregon State University Lines: 26 Message-ID: References: NNTP-Posting-Host: ava.bcc.orst.edu Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII In-Reply-To: On Wed, 13 Mar 1996, C.M.Meghji wrote: > - sorry to hear that you are suffering from no products in your RAPD (NOT > rapid, by the way there is nothing particularly fast about it!). This > phenomenon is the bane of the the RAPD msappers life and it has just > descended on you, but don't feel picked on it happens to EVERYBODY. > The shortest answer is - Discard Everything and start again from > scratch. Your DNA is probably not the problem, get new: water, taq,dNTPs,Mg++ > oil etc. Make up your dNTP,s and whatever in NEW water, and make new DNA diluitions > from your base stock, if you have it. > Replace as much as you can, don't try to eliminate individual elements as the > cause of the problem at this stage. > Good luck, and welcome to the club > > Carol Meghji > ------------------------------------------------------------------------- I did not see the first posting of this message, but I would consider reducing the DNA concentration (e.g. 1 ng/reaction). It's worked for me with DNA extracts from strawberry and hazelnut leaves. Good luck! -Kirk Pomper From owner-rapd@net.bio.net Mon Mar 18 22:00:00 1996 Path: biosci!esvax.dnet.dupont.com!rafalski From: rafalski@esvax.dnet.dupont.com Newsgroups: bionet.molbio.rapd Subject: AmpliTaq vs. Stoffel Date: 19 Mar 1996 13:59:23 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 8 Sender: daemon@net.bio.net Distribution: world Message-ID: <9603192155.AA13822@esds01.es.dupont.com> NNTP-Posting-Host: net.bio.net We have done the experiments showing difference in band size. The reference is: Rafalski, Tingey and Williams Plant Mol. Biol.Manual Supplement H4 pp.1-8 (1994). Kluwer Academic press. See fig.1B (nice picture showing differences in band size). Antoni Rafalski From owner-rapd@net.bio.net Mon Mar 18 22:00:00 1996 Path: biosci!esvax.dnet.dupont.com!rafalski From: rafalski@esvax.dnet.dupont.com Newsgroups: bionet.molbio.rapd Subject: RE: needing to more explanations Date: 19 Mar 1996 14:16:07 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 15 Sender: daemon@net.bio.net Distribution: world Message-ID: <9603192212.AA15516@esds01.es.dupont.com> NNTP-Posting-Host: net.bio.net Your explanation is certainly incorrect. Any shared characters (including RAPD bands) can be used for phylogenetic inference, as long as reproducibility can be established. See for example Tibayrenc, M. Neubauer, K. Barnabe, C. Guerrini, F. Skarecky, D. Ayala, F.J. PNAS 90 (1993) 1335-1339 Genetic characterization of six parazitic protozoa: Parity between random-primer DNA typing and multilocus enzyme electrophoresis Antoni Rafalski From owner-rapd@net.bio.net Mon Mar 18 22:00:00 1996 Path: biosci!RS6000.CMP.ILSTU.EDU!jbcalos From: jbcalos@RS6000.CMP.ILSTU.EDU (Jonathan B. Calos) Newsgroups: bionet.molbio.rapd Subject: listserv address Date: 19 Mar 1996 06:51:15 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: <9603191448.AA40130@rs6000.cmp.ilstu.edu> NNTP-Posting-Host: net.bio.net Desire listserv address to unsubscribe. Thanks. Jon From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!LIFSCI.SDSU.EDU!MCCLELLAND From: MCCLELLAND@LIFSCI.SDSU.EDU Newsgroups: bionet.molbio.rapd Subject: Stoffel versus AmpiTaq Date: 20 Mar 1996 10:07:19 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: <960320100503.20201826@LIFSCI.SDSU.EDU> NNTP-Posting-Host: net.bio.net Sobral and Honeycutt, TAG (1993) were the first to show the utility of Stoffel. These advantages include more bands (of smaller average size) and more primers give good fingerprints. Michael From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!CCC-S.CEDARCREST.EDU!TJLITZI From: TJLITZI@CCC-S.CEDARCREST.EDU (Litzi, Tracy J) Newsgroups: bionet.molbio.rapd Subject: why do RAPD instead RFLP? Date: 19 Mar 1996 21:01:48 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 5 Sender: daemon@net.bio.net Distribution: world Message-ID: <3D8DB05001C93A7C@-SMF-> References: <3D8DB05002C93A7C@-SMF-> NNTP-Posting-Host: net.bio.net What are the benfits of doing RAPD instead of RFLP and are the results obtained from each comparable? From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!galaxy.ucr.edu!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!news.uoknor.edu!news.nodak.edu!plains!comstock From: comstock@plains.nodak.edu (Clay Comstock) Newsgroups: bionet.molbio.rapd Subject: RAPD Primers Date: 20 Mar 1996 01:04:14 GMT Organization: North Dakota Higher Education Computing Network (NDHECN) Lines: 26 Message-ID: <4inlie$lej@daily-planet.nodak.edu> NNTP-Posting-Host: plains.nodak.edu Mime-Version: 1.0 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit X-Newsreader: TIN [version 1.2 PL2] Hello, I am currently an Undergrad at Dickinson State University and I'm doing a project involving RAPD analysis on spiders. I was wondering if anyone had some current information on primers from the University of British Columbia. Last summer I tested set #8 and had good results. If anyone has any input on primers that work for spiders or a similar critters I would be glad to hear from you. Thank You ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Clay Comstock e-mail comstock@plains.nodak.edu homepage http://murphy217.math.dsu.nodak.edu/dsu/comstock.htm PGP Public Key available through most key servers. ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!labatt.com!rob.stewart From: rob.stewart@labatt.com ("Stewart, Robert") Newsgroups: bionet.molbio.rapd Subject: FW: needing to more explanations Date: 20 Mar 1996 05:34:10 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 46 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net ---------- From: Stewart, Robert Sent: Wednesday, March 20, 1996 8:29AM To: rafalski@esvax.dnet.dupont.com Subject: RE: needing to more explanations Greetings, I agree that shared characteristics may suggest that there is some relationship between organisms. However, different characteristics may not necessarily reflect phylogenetic differences. I don't believe it is valid to say because the patterns lack any shared characteristics that the two organisms are not closely related. For example, two bacteria of the same species could have different RAPD patterns. This of course will depend on the primer that is used. Some primers will obviously be better than others. Cheers RS ---------- From: rafalski@esvax.dnet.dupont.com[SMTP:rafalski@esvax.dnet.dupont.com] Sent: Tuesday, March 19, 1996 5:11PM To: rob.stewart@labatt.com; rob.stewart@labatt.com Subject: RE: needing to more explanations Your explanation is certainly incorrect. Any shared characters (including RAPD bands) can be used for phylogenetic inference, as long as reproducibility can be established. See for example Tibayrenc, M. Neubauer, K. Barnabe, C. Guerrini, F. Skarecky, D. Ayala, F.J. PNAS 90 (1993) 1335-1339 Genetic characterization of six parazitic protozoa: Parity between random-primer DNA typing and multilocus enzyme electrophoresis Antoni Rafalski From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!rutgers!uwm.edu!fnnews.fnal.gov!nntp-server.caltech.edu!news.pomona.edu!usenet From: Mira Liao Newsgroups: bionet.molbio.rapd Subject: rapd and molluscs Date: Tue, 19 Mar 1996 21:12:15 -0800 Organization: Pomona College Lines: 8 Message-ID: <314F93AF.59C6@pomona.edu> NNTP-Posting-Host: sclass2.pomona.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (Win16; I) I am working on a senior thesis doing rapd analysis of molluscs. None of my PCR reactions are working, even though I have varied Mg concentration, cycling times, primer concentration, template concentration, etc. etc. I just don't get any amplification at all. I recently read that when extracting mollusc DNA to use for rapd, the phenol-chloroform extraction may not get rid of all of the polysaccharides, leading to poor rapd results. Does anyone know of any other procedure that should be used to extract mollusc DNA in preparation for rapd analysis? From owner-rapd@net.bio.net Tue Mar 19 22:00:00 1996 Path: biosci!ml.csiro.au!chris.bolch From: chris.bolch@ml.csiro.au (Chris Bolch) Newsgroups: bionet.molbio.rapd Subject: Re: FW: needing to more explanations Date: 20 Mar 1996 14:57:10 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 73 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603202253.JAA24480@aqueous.ml.csiro.au> NNTP-Posting-Host: net.bio.net Robert Stewart raises an important point about a lack of shared characters. Almost any phylogenetic analysis requires some yardstick to gauge the relevance/significance of your variation. With a possible exception of rDNA sequences and some mtDNA sequeneces (for which there is a large body of data to assess general levels of divergence at certain taxonomic levels), the significance of the genetic varation uncovered by any method is difficult to assess unless analyses include a range of more and less divergent taxa (i.e outgroups in the analytical sense). This can allow the natural clustering in the analysis to privide some indication of appropriate taxonomic significance. What is more divergent- a single base change in a conserved region of the rDNA, a 10% sequence divergence in an IGS region, or a RAPD genetic distance estimate of 50%?? Hard to say, as we have little understanding of what the 50% RAPD distance means or the relative rates of mutation in many IGS or coding regions in the genome other than rDNA. It easier for those of us who work with morphologically well characterised organisms which have the benefit of complex morphology to provide more charcters for interpreting phylogeny. For those who work with morphologically simple or poorly described/understood groups (such as myself) it is always less obvious what the data means within the context of the existing morphological species concepts. While it may be true that no shared genetic characters does not necessarily mean the organisms are not closely related, if another sample shares more characters then it is a pretty good bet that it is more closely related than the sample which does not. Bacteria, which are notoriously variable within "species", and RAPD (very sensitive to small amounts of sequence variation) are not necessarily a good case in point. If Phylogenetic analysis is the goal, rather than strain discrimination, then the more characters- the better the estimate of phylogeny, therefore use a heap of RAPD primers. Cheers Chris >---------- >From: Stewart, Robert >Sent: Wednesday, March 20, 1996 8:29AM >To: rafalski@esvax.dnet.dupont.com >Subject: RE: needing to more explanations > >Greetings, >I agree that shared characteristics may suggest that there is some >relationship between organisms. However, different characteristics may not >necessarily reflect phylogenetic differences. I don't believe it is valid >to say because the patterns lack any shared characteristics that the two >organisms are not closely related. For example, two bacteria of the same >species could have different RAPD patterns. This of course will depend on >the primer that is used. Some primers will obviously be better than >others. >Cheers >RS ________________________________________ Any views, statements of intent, or comment in this letter have no official status and do not constitute or imply any contract with myself or my employer. ________________________________________ Christopher J. S. Bolch, Algal Ecologist/Geneticist, CSIRO Division of Fisheries, GPO Box 1538, Hobart, Tasmania, Australia, 7001. PH. 002 325314 (Aus), 061-02-325314 (Internat.) FAX. 002 325000 (Aus), 061-02-325000 (Internat.) ________________________________________ From owner-rapd@net.bio.net Wed Mar 20 22:00:00 1996 Path: biosci!ns1.faseb.org!lamarck.sura.net!newsfeed.internetmci.com!howland.reston.ans.net!psinntp!psinntp!psinntp!news.nstn.ca!newsflash.concordia.ca!feed.umontreal.ca!alize.ERE.UMontreal.CA!landryp From: landryp@ERE.UMontreal.CA (Landry Pierre-Alexandre) Newsgroups: bionet.molbio.rapd Subject: Re: RAPD Primers Date: 21 Mar 96 20:59:12 GMT Organization: Universite de Montreal Lines: 36 Distribution: world Message-ID: References: <4inlie$lej@daily-planet.nodak.edu> NNTP-Posting-Host: alize.ere.umontreal.ca comstock@plains.nodak.edu (Clay Comstock) writes: >Hello, >I am currently an Undergrad at Dickinson State University and I'm doing a >project involving RAPD analysis on spiders. >I was wondering if anyone had some current information on primers from >the University of British Columbia. Last summer I tested set #8 and had >good results. >If anyone has any input on primers that work for spiders or a similar >critters I would be glad to hear from you. >Thank You >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ >Clay Comstock >e-mail comstock@plains.nodak.edu >homepage http://murphy217.math.dsu.nodak.edu/dsu/comstock.htm >PGP Public Key available through most key servers. >~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Hello Clay, I have been using set #3 of UBC primers for 7 months now, and had good results for mammals. But since they are the only ones I have tried, I can't compare. Hope this helps. Pierre-Alexandre Landry Dept. des Sciences Biologiques Universite de Montreal C.P. 6128, succursale centre ville Montreal, Quebec H3C 3J7 From owner-rapd@net.bio.net Wed Mar 20 22:00:00 1996 Path: biosci!CS.Arizona.EDU!news.Arizona.EDU!hamblin.math.byu.edu!sol.ctr.columbia.edu!spool.mu.edu!pravda.aa.msen.com!nntp.coast.net!howland.reston.ans.net!vixen.cso.uiuc.edu!usenet.ucs.indiana.edu!willow.bio.indiana.edu!user From: dewolf@bio.indiana.edu (Diana Wolf) Newsgroups: bionet.molbio.rapd Subject: Re: why do RAPD instead RFLP? Date: 21 Mar 1996 00:24:13 GMT Organization: Indiana University Lines: 14 Distribution: world Message-ID: References: <3D8DB05002C93A7C@-SMF-> <3D8DB05001C93A7C@-SMF-> NNTP-Posting-Host: willow.bio.indiana.edu In article <3D8DB05001C93A7C@-SMF->, TJLITZI@CCC-S.CEDARCREST.EDU (Litzi, Tracy J) wrote: > What are the benfits of doing RAPD instead of RFLP and are the results > obtained from each comparable? RAPDs are much faster and easier, and you don't need to radiolable your bands to visualize them as in RFLPs Diana -- Diana Wolf Biology Department Indiana University From owner-rapd@net.bio.net Wed Mar 20 22:00:00 1996 Path: biosci!PUCCINI.CRL.UMN.EDU!martinez From: martinez@PUCCINI.CRL.UMN.EDU ("J. Pat Martinez") Newsgroups: bionet.molbio.rapd Subject: Re: why do RAPD instead RFLP? Date: 21 Mar 1996 08:44:07 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 27 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603211642.KAA10411@puccini.crl.umn.edu> Reply-To: "J. Pat Martinez" NNTP-Posting-Host: net.bio.net Some of the benefits of RAPDs - no need to generate a DNA library - uses about 1000 fold less DNA per run than RFLPs - no radioactive or non-radioactive labeling of nucleic acids - no blotting - quick, once you have the system up and running. Some of the benefits of RFLPs - Markers are more transferable (repeatable) from lab to lab - In my view, are more reliable. - codominance, which allows one to look at Hardy-Weinberg aspects of populations, and provides more information per marker/locus when linkage mapping. > In article <3D8DB05001C93A7C@-SMF->, TJLITZI@CCC-S.CEDARCREST.EDU (Litzi, > Tracy J) wrote: > > > What are the benfits of doing RAPD instead of RFLP and are the results > > obtained from each comparable? % J. Pat Martinez % martinez@puccini.CRL.umn.edu % Univ. of Minnesota % Phone: (612) 625-2221 % Dept. of Plant Pathology % Fax: (612) 625-9728 From owner-rapd@net.bio.net Fri Mar 22 22:00:00 1996 Path: biosci!ns1.faseb.org!lamarck.sura.net!csc.mc.edu!usenet From: "Robert G. Hamilton" Newsgroups: bionet.molbio.rapd Subject: Re: RE: Needing more explanations (RAPDs and phylogenetics) Date: 23 Mar 1996 16:57:09 GMT Organization: Mississippi College Lines: 10 Message-ID: <4j1ah5$2v6@csc.mc.edu> References: <9603192212.AA15516@esds01.es.dupont.com> <4j06aq$eg1@harbour.awod.com> NNTP-Posting-Host: mobo.mc.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Windows; I; 16bit) arthurv@awod.com (Arthur Q. Villordon) wrote: > >a loci present at 344 bp in both samples does not >necessarily mean that these loci share similar homologous sequences >right? Of course this can be proven by taking one step further and use >the markers as probes in hybridization experiments, etc. One must recognize, however, that failure of such a probe to hybridize to RAPDs fragments does not mean the sequences are not homologous. From owner-rapd@net.bio.net Fri Mar 22 22:00:00 1996 Path: biosci!labatt.com!rob.stewart From: rob.stewart@labatt.com ("Stewart, Robert") Newsgroups: bionet.molbio.rapd Subject: Phylogenetics and RAPD Date: 23 Mar 1996 10:05:13 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 19 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Greetings, From the recent discussion of phylogenetics and RAPD, it seems that a couple of things have emerged which would strengthen the use of RAPD for phylogenetic analyses. ( I originally did not agree the use of RAPD and phylogenetics). Michael McClelland pointed out that use of RAPD primers which produce numerous bands, and Chris B. pointed to the need to use a number of different RAPD primers. Both of these are convincing arguments. Thus it seems that if one uses a number of different primers each of which produces numerous bands, then RAPD would yield phylogenetic information. ( An organism which shared 90% of its bands with the test organism would clearly be more closely related than one which shared only 50% of its bands). I wonder, however, how many should be required to make the conclusions from RAPD analyses reliable? Are 5 primers with at least 20 amplified products each sufficient? Would other methods be necessary to validate the conclusions? Cheers RS From owner-rapd@net.bio.net Fri Mar 22 22:00:00 1996 Path: biosci!daresbury!bioftp.unibas.ch!infobiogen.fr!pasteur.fr!univ-lyon1.fr!in2p3.fr!oleane!tank.news.pipex.net!pipex!newsfeed.internetmci.com!battery.awod.com!harbour.awod.com!usenet From: arthurv@awod.com (Arthur Q. Villordon) Newsgroups: bionet.molbio.rapd Subject: RE: Needing more explanations (RAPDs and phylogenetics) Date: 23 Mar 1996 06:39:22 GMT Organization: Ipomoea Lines: 20 Distribution: world Message-ID: <4j06aq$eg1@harbour.awod.com> References: <9603192212.AA15516@esds01.es.dupont.com> NNTP-Posting-Host: chs0092.awod.com Mime-Version: 1.0 Content-Type: Text/Plain; charset=US-ASCII X-Newsreader: WinVN 0.99.7 There certainly is a practical use of RAPD loci in phylogenetic studies, especially in looking at intra-specific relationships. A number of papers have generally "correlated" phylogenetic relationships based on RAPD fragment data vs. morphological and similar data. This assumes of course that shared RAPD loci among samples have homologous or near-homologous sequences, right? However, if inferences (regarding phylogeny) are extended beyond species boundaries, say Ipomoea batatas vs. Ipomoea aquatica, a loci present at 344 bp in both samples does not necessarily mean that these loci share similar homologous sequences right? Of course this can be proven by taking one step further and use the markers as probes in hybridization experiments, etc. Quite a number of papers have expressed concerns regarding inferences made of RAPD loci in inter-specific taxonomic investigations. Arthur ********************************************* From owner-rapd@net.bio.net Sat Mar 23 22:00:00 1996 Path: biosci!grove.iup.edu!YNTHHXA From: YNTHHXA@grove.iup.edu Newsgroups: bionet.molbio.rapd Subject: Primers for Mourning Doves Date: 24 Mar 1996 08:29:08 -0800 Organization: Indiana University of Pennsylvania Lines: 9 Sender: daemon@net.bio.net Distribution: world Message-ID: <01I2PR9LQ76Q8X0DCY@grove.iup.edu> NNTP-Posting-Host: net.bio.net X-URL: mailto:rapd@net.bio.net X-Mailer: Lynx, Version 2-4-1 X-Personal_name: Dawn A. Sherry X-From: YNTHHXA@Grove.iup.edu Hello! I am currently attempting to conduct research on mourning doves (Zenaida macroura). I was wondering if anybody had information on primers that might be appropriate to use with this species. From owner-rapd@net.bio.net Sat Mar 23 22:00:00 1996 Path: biosci!rutgers!csn!news-1.csn.net!imci3!imci2!news.internetMCI.com!newsfeed.internetmci.com!panix!news.eecs.umich.edu!umn.edu!newsstand.tc.umn.edu!menon From: menon@lenti.med.umn.edu (Ravi Menon) Newsgroups: bionet.molbio.rapd Subject: Techniques to study antigenic drift Date: 24 Mar 1996 19:06:35 GMT Organization: University of MN, Medical School Lines: 13 Message-ID: <4j46fr$568@epx.cis.umn.edu> NNTP-Posting-Host: snowman.med.umn.edu X-newsreader: xrn 7.03 Originator: menon@snowman.med.umn.edu We are interested in studying the extent of antigenic drift in a very poorly characterized organism over a two-year period of serial passage in vivo. There is very little known about the surface proteins in this organism, and only two known monoclonal antibodies to surface markers exist. We certainly could use these antibodies to probe the organisms over the course of a number of passages, but if these happen to be against antigenic sites that are invariant, they may not give us much information about the plasticity of surface antigens in general. Are there any obvious ways to tackle this problem, short of putting together RFLPs and RAPDs from square 1? Ravi From owner-rapd@net.bio.net Sun Mar 24 22:00:00 1996 Path: biosci!ml.csiro.au!chris.bolch From: chris.bolch@ml.csiro.au (Chris Bolch) Newsgroups: bionet.molbio.rapd Subject: Re: Phylogenetics and RAPD Date: 24 Mar 1996 16:35:31 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 70 Sender: daemon@net.bio.net Distribution: world Message-ID: <199603250032.LAA12039@aqueous.ml.csiro.au> NNTP-Posting-Host: net.bio.net Dear Rob Stewart You wrote: I wonder, however, how many should be required to make the >conclusions from RAPD analyses reliable? Are 5 primers with at least 20 >amplified products each sufficient? Would other methods be necessary to >validate the conclusions? >Cheers >RS Last question first- I would always try to use a different approach to provide validation or support for the analysis, regardless of which approach I was using. Most sequence studies, for instance, look at one sequence, which is basically only one locus. Conventional wisdom is that for robust, informative phylogenetic studies more loci = better rather than more individuals of each spp. and less loci. The number of bands or primers for reliable analyses is a good question. My approach has been to run 15-20 primers, which usually, under my reaction conditions gave around 300-350 putative RAPD loci. When I analysed the binary scores of my first few datasets I started with 5 primers of data and worked my way upto 15. There came a point (around 10-12) where the phylogenetic reconstructions stabilised. i.e. The branching pattern did not alter with the addition of new data. I take this to be a sign that my data is providing a stable estimate of phylogeny. You could go further and jacknife primers in and out of the analysis to test the robustness of the data set. There are a couple of papers I've seen which have looked at this problem with a few data sets but I can't remember how many primers they recommended or who the authors were. I'd say it depends more on the number of putative loci examined. At some point the "noise" in the dataset will be outweighed by an overall "signal". It will probably vary depending on the reaction conditions and the organisms examined. Also a good question is- How do I select the primers for analysis, and can this bias the data and interpretation? Should you run them randomly and score even those which give poor amplification across your samples, or should you pre-screen for "good primers"? I do the latter so that I reduce the risk of not detecting amplification failure in samples during screening, and for the more bands = better philosophy. Should we select for primers that show strong polymorphism, or select a range or primers which show similarities and differences? Once again I do the latter. Any opinions on these questions would be most welcome. Cheers Chris ________________________________________ Any views, statements of intent, or comment in this letter have no official status and do not constitute or imply any contract with myself or my employer. ________________________________________ Christopher J. S. Bolch, Algal Ecologist/Geneticist, CSIRO Division of Fisheries, GPO Box 1538, Hobart, Tasmania, Australia, 7001. PH. 002 325314 (Aus), 061-02-325314 (Internat.) FAX. 002 325000 (Aus), 061-02-325000 (Internat.) ________________________________________ From owner-rapd@net.bio.net Tue Mar 26 22:00:00 1996 Path: biosci!daresbury!bioftp.unibas.ch!news.vub.ac.be!news.belnet.be!news.rediris.es!scsing.switch.ch!swsbe6.switch.ch!surfnet.nl!howland.reston.ans.net!newsfeed.internetmci.com!in2.uu.net!news1.digital.com!decwrl!news-server.ncren.net!taco.cc.ncsu.edu!news From: Susan Jane Hogarth Newsgroups: bionet.molbio.rapd Subject: Re: Phylogenetics and RAPD Date: Mon, 25 Mar 1996 14:00:31 -0500 Organization: North Carolina State University Lines: 28 Message-ID: <3156ED4F.41C67EA6@unity.ncsu.edu> References: <199603250032.LAA12039@aqueous.ml.csiro.au> NNTP-Posting-Host: fliegen.ent.ncsu.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.0 (X11; I; SunOS 4.1.3_U1 sun4m) Chris Bolch wrote: > Most sequence studies, for instance, look at one > sequence, which is basically only one locus. Really? I've been told that _each_ nucleotide can be treated as one locus. -- Susan Jane Hogarth "Luck is the residue of design." -- Freddy the Fish "Personally, I'm always ready to learn, although I do not always like being taught." -- Winston Churchill http://www4.ncsu.edu/~sjhogart/public/home.html . .-~\ / `-'\.' `- : | / `._ | | .-. { \ | `-' `. . \ | / ~-.`. \| .-~_ `.\-.\ .-~ \ `-'/~~ -.~ / .-~/|`-._ /~~-.~ -- ~ / | \ ~- . _\ From owner-rapd@net.bio.net Tue Mar 26 22:00:00 1996 Path: biosci!RDAVAX.RDA.GO.KR!oh0kw From: oh0kw@RDAVAX.RDA.GO.KR Newsgroups: bionet.molbio.rapd Subject: RAPD in cross-polinated plants Date: 26 Mar 1996 20:32:55 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 3 Sender: daemon@net.bio.net Distribution: world Message-ID: <96032711534001@rdavax.rda.go.kr> NNTP-Posting-Host: net.bio.net I want to try RAPD using chinease bellflower(Platycodon grandiflorum). It's a cross-polinated plant. Is RAPD possible in cross-polinated plant to classify cultivars? From owner-rapd@net.bio.net Sun Mar 31 23:00:00 1996 Path: biosci!UBVMS.CC.BUFFALO.EDU!COFFROTH From: COFFROTH@UBVMS.CC.BUFFALO.EDU (Mary Alice Coffroth) Newsgroups: bionet.molbio.rapd Subject: Acronyms wanted! Date: 1 Apr 1996 12:44:47 -0800 Organization: University at Buffalo Lines: 30 Sender: daemon@net.bio.net Distribution: world Message-ID: <01I316JCZBUG8XH8CJ@ubvms.cc.buffalo.edu> NNTP-Posting-Host: net.bio.net Dear Colleagues, I am putting together a short review of molecular techniques useful in examining population structure for an upcoming symposium. As an introduction I plan to introduce the host of acronyms that are associated with the techniques. Thus I am collecting acronyms and would appreciate hearing from you all. Please include a "translation" and if the technique is new or not wide-spread, a reference would be great, too! Finally, as I hope to introduce as many appropriate techniques as possible, anyone that wants to include their "favorite" approach to population studies would be great. Thanks in advance for any input! By the way, the title for the talk is "Molecular approaches to the study of clonal organisms: Deciphering the alphabet soup". Cheers, Mary Alice Coffroth Department of Biological Sciences State University of New York at Buffalo Buffalo, NY 14260 U.S.A. Email: coffroth@ubvms.cc.buffalo.edu Phone: 716-645-2881 Fax:716-645-2975 * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * *