From owner-rapd@net.bio.net Tue Apr 01 23:00:00 1997 Path: biosci!ACD.TUSK.EDU!Prakash From: Prakash@ACD.TUSK.EDU ("C. S. Prakash") Newsgroups: bionet.molbio.rapd Subject: Large DNA Transfer to Plants Date: 2 Apr 1997 09:39:12 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 77 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Here I reproduce an article I wrote on the Cornell study on transfer of large DNA molecules to plants in the April 1997 issue of ISB NewsReport (http://www.nbiap.vt.edu). *************************************** PLANT GENE TRANSFER COMES OF AGE: LARGE DNA MOLECULES TRANSFERRED TO PLANTS Introduction of foreign DNA molecules into plant cells is a pivotal but routine technique in crop biotechnology. The present technology, however, does not permit the introduction of more than three or four genes at a time. A technique that enables scientists to produce transgenic plants with large inserts of DNA can thus have an exciting impact on both basic and applied biotechnology. Scientists at Cornell University report in the Proceedings of the National Academy of Sciences USA the development of exactly such a technique (1). Using a newly constructed plasmid vector they transferred a 150 kilobase human DNA fragment into plant cells - about ten times the normal DNA size that could be transferred with conventional approaches. The key to the success of the Cornell team, consisting of Carol Hamilton, Steven Tanksley and colleagues, was the methodical development of a unique vector called BIBAC (Binary Bacterial Artificial Chromosome) (2). Libraries with large chromosomal inserts are now increasingly made in the bacterial artificial chromosome (BAC) vectors in E. coli, while the workhorse for transfer of genes into plants is Agrobacterium, which transfers a piece of its DNA into plant cells during infection. The BIBAC vector can multiply in both E. coli and Agrobacterium, and also has additional copies of the virG and virE genes whose products help in the efficient transfer of DNA from Agrobacterium to plant cells. Most of the transgenic tobacco plants developed with the BIBAC vector showed that the introduced human DNA fragment (150 kb) was present in an intact form and the fragment was passed on to subsequent progeny without any changes. Now that the size barrier for plant DNA transfer is broken, scientists can seek to alter quality and yield traits that are controlled by multiple genes. Many agronomically important traits such as seed weight in soybean or tuber dormancy in potato are controlled by such quantitative trait loci. Additionally, many disease resistance genes are known to occur in clusters spanning a large chromosomal segment. The BIBAC vector may, for the first time, facilitate engineering of such complex traits in plants. Tanksley's group has recently isolated a chromosomal segment from tomato containing a major fruit weight quantitative trait locus (3). Richard Michelmore of the University of California, Davis, predicts that BIBAC's most significant applications will be in the map-based cloning of plant genes and a better understanding of how plant genomes are organized (4). According to Hamilton, BIBAC vectors may also be used in the future to reconstitute secondary product pathways, create new pathways for the production of novel compounds and reduce position-dependent expression of transgenes. References 1. Hamilton, C. M., A. Frary, C. Lewis & S.D. Tanksley. 1996. Stable transfer of intact high molecular weight DNA into plant chromosomes. Proc. Natl. Acad. Sci. USA 93:9975-9979. 2. Hamilton, C.M. 1997. Binary-BAC system for plant transformation with high molecular weight DNA. Nuc. Acid Res. (Submitted) 3. Alpert, K.B. & S.D. Tanksley. 1996. High-resolution mapping and isolation of a yeast artificial chromosome contig containing tw2: A major fruit weight quantitative trait locus in tomato. Proc. Natl. Acad. Sci.USA 93:15503-15508. 4. Michelmore, R. 1996. Big news for plant transformation. Nature Biotechnology 14:16553-1654. C. S. Prakash Center for Plant Biotechnology Research Tuskegee University prakash@acd.tusk.edu *************************************** From owner-rapd@net.bio.net Wed Apr 02 23:00:00 1997 Path: biosci!TTACS1.TTU.EDU!brr01 From: brr01@TTACS1.TTU.EDU (Ousama Zaghmout) Newsgroups: bionet.molbio.rapd Subject: (none) Date: 3 Apr 1997 07:47:02 -0800 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 19 Sender: daemon@net.bio.net Distribution: world Message-ID: <01IH9J5ECTAQ96WL20@ttacs.ttu.edu> NNTP-Posting-Host: net.bio.net Please send me information on how to recieve messages from this group. Thanks. Ousama M-Faiz Zaghmout Department of Biological Sciences; Texas Tech University, Lubbock, TX 79409 Phone: 806-742-2709 or 2711 Fax:806-742-2963 E-mail: brr01@ttu.edu Ousama M-Faiz Zaghmout Department of Biological Sciences; Texas Tech University, Lubbock, TX 79409 Phone: 806-742-2709 or 2711 Fax:806-742-2963 E-mail: brr01@ttu.edu From owner-rapd@net.bio.net Sun Apr 06 23:00:00 1997 Path: biosci!agate!spool.mu.edu!uwm.edu!newsfeeds.sol.net!nntp.uio.no!uninett.no!due.unit.no!usenet From: Hans Stenöien Newsgroups: bionet.molbio.rapd Subject: Arlequin ver. 1.0 and RAPD-PCR Date: Mon, 07 Apr 1997 11:00:12 +0200 Organization: The Norwegian University of Science and Technology Lines: 17 Message-ID: <3348B79C.7E6C@vm.unit.no> NNTP-Posting-Host: vm238.ubt.unit.no Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Mailer: Mozilla 3.0Gold (Win95; I) Dear all: I am doing RAPD-PCR analyses on two Sphagnum (peat-moss) species. I am using the haploid gametophores in the analyses. The software Arlequin ver. 1.0 does not accept RAPD-PCR data from diploids. However, shouldn't it be possible to define my data as "RFLP haplotypic data" and carry out ordinary population genetic analyses using this software? Thank you. Sincerely, Hans Stenøien Inst. of Natl. Hist. Museum of Natl. Hist. & Arch. Norwegian University of Science and Technology N-7004 Trondheim, Norway From owner-rapd@net.bio.net Tue Apr 08 23:00:00 1997 Path: biosci!vt.edu!biotech From: biotech@vt.edu (Don Ball, by way of C. S. Prakash) Newsgroups: bionet.molbio.rapd Subject: Biotechnology 2001 Conference for Teachers and Science Educators Date: 9 Apr 1997 16:51:58 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 432 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Biotechology 2001 Conference Updating Educators for the 21st Century June 20-21, 1997 F E A T U R I N G Dr. Roger Beachy, Scripps Oceanographic Institute La Jolla, California Dr. Esther Chang, Georgetown Medical Center Washington D.C. Dr. Scott Woodward, Brigham Young University Salt Lake City, Utah *** SPECIAL LODGING RATE ($36/DAY) AVAILABLE TO EDUCATORS (see details below) *** ALSO SPECIAL FRIDAY MEETING RATES FOR FULLTIME STUDENTS (inquire Dr. Rebecca Ross, 540 231-9088 or rrwings@vt.edu) Sponsored by: Virginia Tech Fralin Biotechnology Center and Division of Continuing Education in collaboration with The National Association of Biology Teachers HIGHLIGHTS Biotechnology 2001 is Virginia's second major biotechnology conference for science educators. The goal of the conference is to help science educators keep up with scientific discoveries and related developments in biotechnology. New discoveries and their application to medicine, agriculture, and the environment are occurring at an astounding pace. Biotechnology will have far reaching implications for almost every aspect of our lives. However, keeping up to date is a daunting task for educators. Therefore Virginia Tech's Fralin Biotechnology Center and the Division of Continuing Education, in collaboration with the National Association of Biology Teachers, have organized Biotechnology 2001. We encourage high school and college faculty to take advantage of this unique opportunity, but educators in all disciplines and other interested persons are also welcome to attend. The first day's program features nationally and internationally recognized scientists. The symposium will be held at the Donaldson Brown Hotel and Conference Center at Virginia Tech, Blacksburg, Virginia, on Friday, June 20. Participants paying full registration fee may visit Virginia Tech's Fralin Biotechnology Center on Saturday, June 21, for lectures by Virginia Tech scientists and tours of biotechnology laboratories, or participate in workshops at the Fralin Center, Engel, or Cheatham laboratories on the Virginia Tech campus. LOCATION AND LODGING The Donaldson Brown Hotel and Conference Center is located on the Virginia Tech campus in Blacksburg, Virginia. The Center is located near downtown, two blocks off Main Street at the corner of College Avenue and Otey Street. Parking permits are required and are available at the Donaldson Brown Hotel lodging desk. A block of lodging rooms has been reserved at the hotel for Thursday, June19, and Friday, June 20. Rate is $62 per night, single or double. To reserve a room, call (540) 231-5156 by May 10. Check here for other accomodations ********** A LIMITED NUMBER OF AIR CONDITIONED DORM ROOMS ARE AVAILABLE. $36 / DAY SINGLE, $32 / DAY DOUBLE. BOTH INCLUDE MEALS (student dining halls) PER DAY. ********** HOW TO REGISTER: THE REGISTRATION FEE FOR THE CONFERENCE IS $110. This fee includes all refreshment breaks, lunch both days, handouts, speakers and workshops. Please complete the attached registration form and return with payment by MAY 10, 1997. AFTER THIS DATE, the registration fee is $130 and should be confirmed by phone with the conference registrar at (540) 231-5182. REFUND AND CANCELLATION POLICY: Requests for refunds are honored if received four full working days prior to the conference. The university may cancel or postpone any course because of insufficient enrollment or other unforeseen circumstances. If the course is cancelled or postponed, the university will refund registration fees, but cannot be held responsible for any other costs, charges, or expenses, including cancellation/change charges assessed by airlines or travel agencies. FOR MORE INFORMATION: For further information about the conference, please contact Rebecca Ross at (540) 231-9088, e-mail at rrwings@vt.edu, or fax at (540) 231-7126. CONFERENCE SCHEDULE: FRIDAY, JUNE 20 (held at The Donaldson Brown Hotel and Conference Center, Blacksburg, Virginia) 7:30-8:30 Coffee and Registration Check-In 8:30-8:45 Welcome, Dr. Tracy Wilkins-Director of The Fralin Biotechnology Center, Virginia Tech, and President of TechLab, Inc., Blacksburg, VA 8:45-9:30 Dr. Roger Beachy--Head, Division of Plant Biology, The Scripps Research and Oceano-graphic Institute, La Jolla, CA, "The Impact of Biotechnology on Agriculture in Developed and Developing Countries" 9:30-10:15 Dr. John Quackenbush-Director of Full Length cDNA Sequencing Project, Institute for Genomic Research, Rockville, MD,"The Once and Future Genome Project"-Mapping, Sequencing, and Ethical Issues of the HGP 10:15-10:45 Break 10:45-11:30 Dr. Betty Mansfield-Editor of the Human Genome News, Oak Ridge National Laboratories, Oak Ridge, TN, "The Human Genome Project: Fueling Research Into The Next Millennium" 11:30-12:15 Dr. Esther Chang-Researcher, Lombardi Cancer Center, Georgetown University Medical Center, Washington, D.C.,"The p53 Tumor Suppressor Gene: How Is It Being Used?" 12:15-1:30 Buffet Luncheon 1:30-2:15 Dr. Tracy Wilkins-Director of Fralin Biotechnology Center, Virginia Tech, and President of TechLab, Inc., Blacksburg, VA,"Transgenic Plant and Animal Research: Generating New Products for the 21st Century" 2:15-3:00 Mr. Larry Pressley-Special Agent, Federal Bureau of Investigation Laboratories, Quantico, VA,"Quality Assurance and Forensic DNA Testing in Criminal Investigations" 3:00-3:30 Break 3:30-4:15 Dr. Ilya Raskin-AGBIOTECH Center, Rutgers University, New Brunswick, NJ,"Genetically Engineered Plants for Bioremediation of the Environment" 4:15-5:00 Dr. Joseph Falkinham-Biology Department and The Fralin Biotechnology Center, Virginia Tech, Blacksburg, VA,"DNA Fingerprinting: Identifying Sources of Bacterial Infection in AIDS Patients" 5:00-5:30 Break 5:30-6:30 Dr. Scott Woodward-Microbiology Department and Head of Egyptian DNA Project, Brigham Young University, Salt Lake City, UT, "Ancient DNA Studies of the 18th Egyptian Dynasty Pharoahs" WORKSHOPS and FIELDTRIPS SATURDAY, JUNE 21 ADVANCE REGISTRATION (by MAY 10) REQUIRED FOR ALL Workshops held at Fralin, Engel, or Cheatham labs on campus 1. Seafood Forensics: Electrophoresis of Proteins Using Agarose Gels, Dr. Jonathan Morris, Biotechnology Program Coordinator, Middlesex Community College, Middletown, CT, and Mr. Harry Dammers, Glastonbury High, Glastonbury, CT. Time: 9:00am-12:00pm/ Limit: 24 This lab workshop will familiarize the participant with Protein Electrophoresis techniques using horizontal agarose gels and forensic analysis of crab tissue to determine if species substitution has occurred. This lab is part of the NABT/LTI High Quality Biotechnology 'On A Shoestring' Program. Partial support provided by NSF/ATE Program Grant #DUE 9553720 Location: Virginia Tech Campus 2. DNA Sequencing Using a Paper and Paper Clip Simulation, Dr. Helen Kreuzer of Carolina Biological Supply Co., and one of the authors of Recombinant DNA and Biotechnology, A Guide for Teachers, published by American Society for Microbiology Time: 10:00am-12:00pm/Limit: 24 In this hands-on activity the concept of the dideoxy sequencing method will be illustrated using paper templates and colored paper clips. Concepts of how chain terminators work and how they can be used to find the base sequence of a DNA molecule will be revealed by pooling results of the group, symbolically running them in a sequence gel, and then reading them. Actual autoradiographs will be examined as well. Location: Virginia Tech Campus 3. The Human Genome Project: Biology, Computers, and Privacy (A Computer Instructional Module Developed By BSCS and Supported by the Department of Energy). Mr. Joe McInerney, Director of BSCS, Colorado Springs, CO Time: 8:00-10:00am/Limit: 24 Mr. McInerney, one of the developers of this computer module, will take you through selected activities of the module in which students learn about the uses, limitations, and implications of genomic databases. The module provides teachers with information on the HGP and the ethical, legal, and social issues related to genomic databases. Mr. McInerney will give you a free copy of this module and also tell you about BSCS's third genome module released in 1997. Location: Virginia Tech Campus 4. Genes and Cancer (All cancer is genetic! Some cancers are inherited!), Mr. Sam Rhine, Director of the Genetic Ed Center, Fortville, IN Time: 9:00am-12:00pm/Limit: none This lecture/slide presentation is a thought-provoking look at the activation and conversion of proto-oncogenes into oncogenes; the inactivation of tumor suppressor genes; apoptosis; mismatch repair; tumor evolution; and gene therapies for cancer. Mr. Rhine will have tapes and fifty page outline booklets available for purchase. Location: Virginia Tech Campus 5. Secrets of the Rain Forest, Mr. Ron Mardigian, developer for BIO-RAD Laboratories, Hercules, CA *Major Cost Underwritten by BIO-RAD Laboratories Time: 9:00am-12:00pm/Limit: 60 The goal of this new experimental kit is for participants to isolate and clone a hypothetical stomach cancer curing protein (GFP) produced in the leaves of a tree that grows in the Andean Rain Forest. All of the genes from the leaves have been cut and pasted into cloned bacterial cells. In order to find the bacteria containing the therapeutic protein the bacteria are streaked onto selective media. All colonies appear white under normal room light, but under UV light, colonies containing the cancer curing gene appear bright green. The green colonies are then picked off the agar and grown overnight in liquid media. The bacteria are then lysed to release the GFP and the desired product is purified from contaminating bacterial proteins by passage over a hydrophobic interaction chromatography column. This new kit and two week curriculum developed by BIO-RAD incorporates transformation and purification techniques employed by biotech companies to get a product to market, as well as ethical, economic, and regulatory procedures that must be adhered to. Location: Virginia Tech Campus 6. Protein Purification Methods: Column Chromatography, Dr. Kristi DeCourcy, Fralin Biotechnology Center, Virginia Tech, Blacksburg, VA, and Mrs. Ellen Mayo, Mills Godwin High School, Richmond, VA Time: 9:00am-12:00pm/Limit: 16 Participants will learn to do column chromatography, ion-exchange chromatography, and reverse-phase chromatography of proteins. These methods are used by genetic engineering companies to isolate and purify products such as insulin, interferons, and TPA expressed by genetically engineered organisms. Participants will pour their own columns and also use pre-packed columns. Location: Virginia Tech Campus 7. Using Electrophoresis in a PCR Forensics Simulation, Mr. W. Todd Grey, of Carolina Biological Supply Co., Burlington, NC Time: 10:00am-12:00pm/Limit: 24 *Major Cost Underwritten by Carolina Biological Supply Company PCR amplification of certain highly variable human alleles often results in different electrophoresis patterns. When multiple alleles obtained from a person are amplified and electrophoresed, a unique pattern, known as a DNA fingerprint, is produced. In this workshop, participants will electrophorese DNA that simulates samples obtained by PCR and compare patterns of two "suspects" to patterns of DNA obtained at a crime scene. Location: Virginia Tech Campus 8. Talks and Tours at Fralin Biotech Center Research Labs, Virginia Tech, Blacksburg, VA Includes lunch/ Limit: 60 Time: 8:30-10:30am-Tours-Meet in Auditorium of Fralin Center-First Floor 1. Labs of Dr. Carole Cramer and Dr. Beth Grabau-Transgenic Tobacco and Soybeans 2. Lab of Dr. Joe Falkinham- Mycobacterium tuberculosis and AIDS Research 3. Lab of Dr. Dennis Dean-Nitrogen Fixation 4. Lab of Dr. Nancy Love-Environmental Biosensors and Remediation 5. Lab of Dr. Richard Helm-Wood Chemistry 6. Lab of Dr. Tracy Wilkins-Microbial Toxins Time: 10:30-11:30am-Lecture-Fralin Auditorium-First Floor Dr. Sue Tolin-Plant Pathology Department, Virginia Tech, "Finding Genes To Protect Plants From Viruses" Time: 11:30am-12:00pm-Computer Presentation-Fralin Auditorium-First Floor Mr. Doug King-Biochemistry Department, Virginia Tech, "Accessing the AgBiotech Homepage Can Help You!" What's the latest on plant pesticide genes, edible plant vaccines, altering mosquitoes so that they no longer act as malaria vectors? Find out how you can access this information and more. 9. Using Computers To Improve Your AP Biology Labs, Dr. Charles Lytle,Coordinator of Biology Outreach Programs, North Carolina State University and Former Chair of National AP Biology Committee, Ms. Bobbie Hinson, Providence Day School, Charlotte, NC, and Ms. Judy Powell, Coordinator, Howard Hughes Precollege Outreach Program and The Science House, North Carolina State University. Time: 1:00-5:00pm/Limit: 20 Most of the investigative laboratory exercises recommended for Advanced Placement Biology are easily adaptable to computer-based labs using appropriate computer probes and software. Teachers will learn to use the software and probes to provide new opportunities for students to get practice in scientific inquiry and to learn the value of computer technology in research. Location: Virginia Tech Campus 10. Introductory Cell and Tissue Culture Techniques and Karyotyping of Human Chromosomes, Dr. Roland Nardone, Director of the Discovery Center for Cell and Molecular Biology, The Catholic University of America, Washington, D.C. Time: 1:00-5:00pm/Limit: 20 The cell and tissue culture-biotechnology interface is very broad and includes growth of cells for valuable products, gene therapy, and production of hybridomas for monoclonal antibody production. Karyotyping analysis is a core element of the human genome project which endeavors to locate the 50,000-100,000 genes of the chromosomes and also has application in research of genetic diseases, cancer, gene expression, evolution, and mutagenicity. Participants will obtain hands-on experience in the key procedures of cell and tissue culture and karotyping. Information packets including lecture notes and lab protocols will facilitate information transfer. Location: Virginia Tech Campus 11. The Human Genome Project: Biology, Computers, and Privacy (A Computer Instructional Module Developed By BSCS and Supported by the Department of Energy), Mr. Joe McInerney, Director of BSCS, Colorado Springs, CO Time: 1:00-3:00pm/Limit: 24 Mr. McInerney, one of the developers of this computer module, will take you through selected activities of the module in which students learn about the uses, limitations, and implications of genomic databases. The module provides teachers with information on the HGP and the ethical, legal, and social issues related to genomic databases. Mr. McInerney will give you a free copy of the module and also tell you about BSCS's third genome module released in 1997. Location: Virginia Tech Campus 12. The Access Excellence Program-A Resource for Labs and Information, Access Excellence Fellows Mrs. Ellen Mayo, Mills Godwin High School, Richmond, VA, Dr. Leslie Pierce, Thomas Edison High School, Alexandria, VA, and Mrs. Barbara Kolb, James River High School, Buchanan, VA Time: 3:30-4:30pm/Limit: 50 How to access via internet this program developed by Genentech, Inc., and download lab activities, talk to colleagues, talk to bench scientists, and keep abreast of cutting-edge biotech information via your computer. Visit at http://www.gene.com/ae Location: Virginia Tech Campus 13. DNA Sequencing Using a Paper and Paper Clip Simulation, Dr. Helen Kreuzer, of Carolina Biological Supply Co., and one of the authors of Recombinant DNA and Biotechnology, A Guide for Teachers, published by American Society for Microbiology Time: 3:00-5:00pm/Limit: 24 In this hands-on activity the concept of the dideoxy sequencing method will be illustrated using paper templates and colored paper clips. Concepts of how chain terminators work and how they can be used to find the base sequence of a DNA molecule will be revealed by pooling results of the group, symbolically running them in a sequence gel, and then reading them. Actual autoradiographs will be examined as well. Location: Virginia Tech Campus 14. What's New In Genetics, Cytogenetics, and Reproductive Biology?, Mr. Sam Rhine, Director of the Genetic Ed Center, Fortville, IN Time: 1:00-3:00pm/Limit: none This lecture/slide presentation will overview some of the new techniques and current topics being discussed in the literature such as Banding, FISH, Painting and Microdissection, Karyotyping, Colored Karyotypes, Chromosome syndromes, X-Inactivation, the Lyon Hypothesis, the HGP, RFLPs and their applications, VNTRs, STRPs, etc. Location: Virginia Tech Campus 15. Secrets of the Rain Forest, Mr. Ron Mardigian, developer for BIO-RAD Laboratories, Hercules, CA *Major Cost Underwritten By BIO-RAD Laboratories Time: 2:00-5:00pm/Limit: 60 The goal of this new experimental kit is for participants to isolate and clone a hypothetical stomach cancer curing protein (GFP) produced in the leaves of a tree that grows in the Andean Rain Forest. All of the genes from the leaves have been cut and pasted into cloned bacterial cells. In order to find the bacteria containing the therapeutic protein the bacteria are streaked onto selective media. All colonies appear white under normal room light, but under UV light, colonies containing the cancer curing gene appear bright green. The green colonies are then picked off the agar and grown overnight in liquid media. The bacteria are then lysed to release the GFP and the desired product is purified from contaminating bacterial proteins by passage over a hydrophobic interaction chromatography column. This new kit and two week curriculum developed by BIO-RAD incorporates transformation and purification techniques employed by biotech companies to get a product to market, as well as ethical, economic, and regulatory procedures that must be adhered to. Location: Virginia Tech Campus 16. High School and Community College Genome Program, Dr. Maureen Munn, University of Washington, Seattle, WA, and Ms. Diane Lashinsky, Shorecrest High, Seattle, WA Time: 8:30am-5:30pm/Limit: 20 This all-day workshop is targeted for high school teachers, community college faculty, and scientists who are interested in developing sequencing programs in their communities. Participants will sequence DNA using classroom-friendly techniques and will discuss the many ethical issues involved in pre-symptomatic testing for Huntington's disease. The program will cover teaching strategies, integration of the material into the curriculum, and ideas for the development and implementation of a teacher-scientist partnership for a sequencing program. Location: Virginia Tech Campus 17. DNA Amplification by Polymerase Chain Reaction/Simulation of the p53 Tumor Suppressor Gene/Simulation of ELISA to Detect AIDS (Advanced)/College, Dr. Jack Chirikjian, Department of Biochemistry and Molecular Biology, Georgetown University School of Medicine, Washington, D.C. Time: 9:00am-4:00pm/Limit: 24 In this all-day workshop, participants will work through three "cutting edge" laboratory exercises: First, the polymerase chain reaction will be used to amplify a DNA fragment using a set of two primers, one for each DNA strand. A thermal cycler will be used to create the temperature oscillations required. The second exercise will use a new kit developed by EDVOTEK which simulates the p53 gene and its mechanisms for suppression of tumor growth. The third lab exercise will illustrate metho-dologies involved with enzyme-linked immunosorbent assay (ELISA) through a simulation of a clinical screening of serum samples for the presence of antibodies for the AIDS virus. (The experiment does not use any human or blood products.) Location: Virginia Tech Campus *Virginia Tech does not discriminate against employees, students, or applicants on the basis of race, sex, handicap, age, veteran status, national origin, religion, political affiliation, or sexual orientation. Anyone having questions concerning discrimination should contact the Equal Opportunity/Affirmative Action Office. If you are a person with a disability and require any auxiliary aids, services, or other accommodations for this workshop, please discuss your accommodation needs with Barbara Duncan at (540) 231-4849 at your earliest convenience. _________________________________ Webmaster: biotechweb@vt.edu (Bob Mock) WWW: http://www.biotech.vt.edu/ Don Ball Fralin Biotechnology Center Virginia Polytechnic Institute and State University Blacksburg, VA 24061-0346 USA 540 231-6934 540 231-7126 fax biotech@vt.edu From owner-rapd@net.bio.net Tue Apr 08 23:00:00 1997 Path: biosci!daresbury!not-for-mail From: Roland Koelliker Newsgroups: bionet.molbio.rapd Subject: RAPD with Orchard Grass Date: 9 Apr 1997 13:39:17 +0100 Lines: 28 Sender: lpddist@mserv1.dl.ac.uk Distribution: bionet Message-ID: <5ig2ll$17p@mserv1.dl.ac.uk> X-Sender: rkoellik@weizen.ethz.ch Original-To: rapd@dl.ac.uk Dear netters I have RAPD's working very well with perennial ryegrass (Lolium perenne) and meadow fescue (Festuca pratensis). Now I'd like to investigate orchardgrass (dactylis glomerata). I tried several primers that worked for the other species, but I did never get any amplification. Does anybody have an idea what the problem could be? Thank you for your help. ************************************************ !!!!! New mail address: roland.koelliker@ipw.agrl.ethz.ch !!!!!! ************************************************ *************************** Roland Koelliker Swiss Fed Inst Technol Plant Sciences ETH-Zentrum, LFW-C36.1 CH-8092 Zurich Tel ++41-1-632 3888 Fax ++41-1-632 1153 e-mail: roland.koelliker@ipw.agrl.ethz.ch *************************** From owner-rapd@net.bio.net Sat Apr 12 23:00:00 1997 Path: biosci!fcs280s.ncifcrf.gov!cpk-news-feed1.bbnplanet.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!feed1.news.erols.com!insync!uunet!in3.uu.net!206.67.134.2!hst1us.satlink.com!satlink!un3.satlink.com!news From: "Estela Martinez y Manuel Criado" Newsgroups: bionet.molbio.rapd Subject: rapd Date: 13 Apr 1997 13:30:34 GMT Organization: Satlink, Argentina Lines: 7 Message-ID: <01bc4827$e46ffc40$1cea00c8@default> NNTP-Posting-Host: ppp-22.mza.satlink.com X-Newsreader: Microsoft Internet News 4.70.1155 Hello I would like to enter to this group and to know about the problem of the RAPD. Thank you. Liliana From owner-rapd@net.bio.net Sun Apr 13 23:00:00 1997 Path: biosci!byu.edu!JLFarmer From: JLFarmer@byu.edu (James Farmer) Newsgroups: bionet.molbio.rapd Subject: RE: Help with a Posting Date: 14 Apr 1997 15:59:42 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 142 Sender: daemon@net.bio.net Distribution: world Message-ID: <3.0.1.32.19970414165914.006a01e0@apollo.byu.edu> NNTP-Posting-Host: net.bio.net This looks like a legitimate posting to me, even though it is peripheral to our major concerns. To those of you who are not US citizens, please accept my apology and press the delete button. James Farmer >Return-path: >Date: Mon, 14 Apr 1997 17:26:19 +0000 >From: "Robert W. Georgantas III" >Subject: Help with a Posting >Sender: rgeorgan@welchlink.welch.jhu.edu >To: RAPD James Farmer > >To: James Farmer >Moderator of the RAPD BioNet Discussion Group. > >I was hoping that you could help me, please read the letter below then do >with it as you wish. > >I am a student with the Department of Pharmacology and Molecular Sciences >at Johns Hopkins University, School of Medicine. I have started up a >small internet site called "The Science Guide" located at the URL >http://www.scienceguide.com. I started the site to address the lack of a >unifying source of information for scientists on the internet. As you >can see in the announcement below the Guide consists of a number of >sections designed to help scientists find information on the internet, >The news section is currently our most popular service receiving over 4K >visits per day, with a hundreds of subscribers to the Daily News Emailer. > >Since we do not charge for our service, and since we are not accepting >advertisers; we have only a small number of routes by which to spread the >word about the Guide (we have no advertising budget as well). One of >these routes is by posting to Usenet groups. We have done this on a >small basis by looking at a news group to assure that the users of the >group are appropriate for our service, and then posting the Announcement >enclosed below to the group. We Do Not Spam. Over the last month we >have posted to roughly forty news groups and have received a great >response in the form of emails telling us how useful our site is, and >will become as we add more content to it. We have not received a single >negative response, which we have interpreted to mean that our site is >useful enough to scientists that they do not mind our "off-subject" >posting. > >That brings me to the crux of the email, I was hoping that you could take >the time to visit the Science Guide, determine if it would be of use to >the subscribers of the group which you modify, and if so - post the >announcement text below to the group. This would be of great help to us >in getting the word out about the Guide. > >If you feel that this posting would be inappropriate, please accept my >apologies for having taken your time, and just disregard this letter. > >One thing that may be particularly of interest, is our plan to start >monitoring Congress for bills containing items concerning scientific >funding. We plan on doing this to take advantage of the power of the >internet to set up a pseudo lobbying group. As an example - during the >Congressional Debates over the NIH budget a few months ago we encouraged >our readers to email their congress-person asking for an increase in the >budget. At the time we had just started the guide and had only a small >number of readers, but now that we have literally thousands of users (and >our usership is growing at 40% per week) and we think that we could make >a significant difference in funding by informing our users of congresses' >actions and helping them to act in writing to the House, or Senate, or >President. > >Thank you >Robert Georgantas - The Science Guide > >PS. You can check to make sure that I am actually who I say that I am >buy fingering my student email account: rgeorgan@welchlink.welch.jhu.edu. > The Science Guide is hosted under the .com domain by Hway Technologies, >because Hopkins was not willing to host a site with our potential traffic. > > >------------------Announcement Text------------------------ > >Announcing the SCIENCE GUIDE. >http://www.scienceguide.com > >A New Internet Directory and Information Service run by Scientists and >Physicians for Scientists and Physicians. After visiting the Guide, If >you have any suggestion for making the Guide better please let us know. >(webmaster@scienceguide.com) > >The Science Guide consists of a number of different sections designed to >help the scientist and physician find information on the internet and to >sponsor communication between those interested in science: > > >NEWS SECTION > >Every day the Science Guide compiles medical and research news from >national news sources around the net. Most of the news articles are >concerned with medicine, bioscience, and physics, but all other sciences >from agriculture to zoology are commonly included. News sources currently >listed include: CNN, EurekAlert, HMS Beagle, MSNBC Sci-Tech, Science >Magazine's ScienceNow, CBS Space News, USA Today, The Albuquerque >Journal, Scientific American Web Weekly, The Why Files, Discover >Magazine, Scientific American, Smithsonian Magazine, and the Technology >Review. The news pages also list links to news sources not compiled >within the News site. We are currently working on adding a number of >other sources to the site to make it even more useful. > >To make getting science news even easier, we send out a DAILY NEWS >EMAILER listing the articles which have been compiled on our site. >Anyone can subscribe to the Emailer by sending an email to >news@scienceguide.com with the message "Subscribe" > > >DIRECTORY OF USENET NEWS GROUPS and DISCUSSION LISTS > >The Directory of Usenet and Discussion Groups is compiled quarterly from >different sources around the net to provide the scientist and those >interested in science easy access to these invaluable sources of >discourse and information. We are currently working on finding the >proper subscription method for each of the discussion lists. This is >taking a bit longer that we thought so please pardon our dust. The >Usenet portions of this section are complete. > > >ON-LINE JOURNAL HYPERLINK SECTION > >The Journals Section contains links to peer reviewed scientific journals >on the Internet. Each listing clearly indicates whether the journal >provides only the table of contents, TOC with abstracts, or the full text >of the journal > > >EMPLOYMENT SECTION > >The Jobs and Positions Section contains hyperlinks to the best Scientific >Employment Databases and Classifieds on the net. > > >GRANTS and FUNDING SECTION > >The funding section contains links to the best funding and grant >databases on the Internet, making it very easy for scientists to quickly >find funding opportunities. The featured site of the section is "The >Community of Science," a Johns Hopkins service designed to help >scientists find and continue funding. > From owner-rapd@net.bio.net Mon Apr 14 23:00:00 1997 Path: biosci!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!feed1.news.erols.com!news.maxwell.syr.edu!worldnet.att.net!howland.erols.net!vixen.cso.uiuc.edu!news.indiana.edu!willow.bio.indiana.edu!user From: dewolf@bio.indiana.edu (Diana Wolf) Newsgroups: bionet.molbio.rapd Subject: Re: RAPD with Orchard Grass Date: 15 Apr 1997 22:38:30 GMT Organization: Indiana University Lines: 23 Distribution: bionet Message-ID: References: <5ig2ll$17p@mserv1.dl.ac.uk> NNTP-Posting-Host: willow.bio.indiana.edu In article <5ig2ll$17p@mserv1.dl.ac.uk>, Roland Koelliker wrote: > Dear netters > > I have RAPD's working very well with perennial ryegrass (Lolium perenne) > and meadow fescue (Festuca pratensis). Now I'd like to investigate > orchardgrass (dactylis glomerata). I tried several primers that worked for > the other species, but I did never get any amplification. Does anybody have > an idea what the problem could be? > > Thank you for your help. > Perhaps your orchardgrass DNA is dirty or too concentrated. Are you cleaning your DNA? Are you checking DNA concentrations? I would try diluting the DNA alot to see if it will amplify. Diana -- Diana Wolf Biology Department Indiana University From owner-rapd@net.bio.net Wed Apr 16 23:00:00 1997 Path: biosci!greb.ulaval.ca!Christian.Mouton From: Christian.Mouton@greb.ulaval.ca (Christian Mouton) Newsgroups: bionet.molbio.rapd Subject: Re: Looking for primer design program Date: 17 Apr 1997 07:04:51 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 36 Sender: daemon@net.bio.net Distribution: world Message-ID: <199704171407.KAA26323@hermes.ulaval.ca> NNTP-Posting-Host: net.bio.net >Dear Netters, >I am trying to design specific primers about 21 bps according to the sequen= ces >of polymorphic bands from RAPD reactions. Does anyone know a good >computer program to do that on a PC? The demo version of Primer Premier 4 >looks fine except that it costs $485US (basic version). I have been using = the >Oligos 3 program which is quit old and I have to select the probable primer >sequence manually. I have more than 30 of the polymorphic RAPDs to convert >to SCARs so I am really looking for a some what automatic way to do that. > Dear Kae-kang Hwu: I believe that you should not expect any software to give you right away the perfect primers and perfect amplification conditions. My lab has some experence in the design of specific primers using OLIGO TM Primer Analysis Software version 5.0 as described in Guillot & Mouton 1996 Molecular and Cellular Probes 10:413-421. In each case we had to work around the predicted sequences. In particular, for each potentially suitable primer pair the optimal annealing temperature was determined experimentally. A series of experiments were performed in which annealing temperatures were adjusted degree by degree within a range that extends from -5=B0C to +15=B0C from the temperature predicted with the help of the software. I hope this will help. Christian Mouton, DCD, DSO Groupe de Recherche en Ecologie Buccale =46aculte de medecine dentaire, Universite Laval Quebec (Quebec) G1K 7P4 CANADA tel. (418)656-5872; fax. (418)656-2861 From owner-rapd@net.bio.net Wed Apr 16 23:00:00 1997 Path: biosci!CCMS.NTU.EDU.TW!khwu From: khwu@CCMS.NTU.EDU.TW (Kae-kang Hwu) Newsgroups: bionet.molbio.rapd Subject: Looking for primer design program Date: 16 Apr 1997 17:35:23 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 14 Sender: daemon@net.bio.net Distribution: world Message-ID: <1.5.4.32.19970417003353.006953d4@ccms.ntu.edu.tw> NNTP-Posting-Host: net.bio.net Dear Netters, I am trying to design specific primers about 21 bps according to the sequences of polymorphic bands from RAPD reactions. Does anyone know a good computer program to do that on a PC? The demo version of Primer Premier 4 looks fine except that it costs $485US (basic version). I have been using the Oligos 3 program which is quit old and I have to select the probable primer sequence manually. I have more than 30 of the polymorphic RAPDs to convert to SCARs so I am really looking for a some what automatic way to do that. Kae-kang Hwu Dept. of Agronomy, National Taiwan University From owner-rapd@net.bio.net Wed Apr 16 23:00:00 1997 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.rapd Subject: BIOSCI/bionet miniFAQ & Fundraiser Date: 17 Apr 1997 02:00:10 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 239 Sender: daemon@net.bio.net Distribution: world Message-ID: <199704170900.CAA01609@net.bio.net> NNTP-Posting-Host: net.bio.net (LAST REVISION: 30-JUL-95) This BIOSCI "miniFAQ" is designed to answer the questions that come up the *most frequently*. The main BIOSCI FAQ (Frequently Asked Questions) is accessible on the World Wide Web at URL http://www.bio.net/. If you can not find an answer to your question in this or other documentation, the BIOSCI technical support staff answers e-mail queries sent to biosci-help@net.bio.net We can only answer questions about the use of the newsgroups and mailing lists. We unfortunately do not have the staff to do Internet information searches or answer scientific questions. Please post those to the appropriate BIOSCI/bionet newsgroups. Contents: -------- 0) BIOSCI NEEDS YOUR SUPPORT!! 1) Using the WWW to access the BIOSCI/bionet newsgroups. 2) What to do about "spams," i.e., junk mail, ads, etc. 3) Examples of subscribing and unsubscribing to the mailing lists. 4) The BIOSCI user address and research interest directory. 0) BIOSCI NEEDS YOUR SUPPORT!! ------------------------------ BIOSCI's government funding has been expended, and we are now operating solely from advertising revenue that we have raised from our Web site at http://www.bio.net/. We need just a few minutes of your time to help us serve you. You can do two important things which will take very little time for you individually and will immensely help us continue to help you. First, please use our WWW system at http://www.bio.net/ to access the archives. You can post or reply to messages via your Web browser as described in item #1 below. 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. 1) Using the WWW to access the BIOSCI/bionet newsgroups. -------------------------------------------------------- As of 10 December 1995, all BIOSCI/bionet full newsgroups are accessible through the World Wide Web (WWW) at URL http://www.bio.net. One can read and reply publicly or privately to both recent postings and archived messages through one's Web browser if it is configured properly to send e-mail. Each newsgroup is equipped with its own WAIS index. The main BIOSCI home page also has access to the BIO-JOURNALS Table of Contents database WAIS index and the BIOSCI user address database described in another item further below. 2) What to do about "spams," i.e., junk mail, ads, etc. ------------------------------------------------------- BIOSCI is a set of parallel USENET newsgroups (the "bionet" groups), mailing lists, and a hypermail archive at URL http://www.bio.net/. The same postings are distributed on all media (except for a small number of mailing-list-only groups at net.bio.net). Unfortunately it is becoming a despicable practice on the Internet (by a few people out to make a fast buck) to do automated mass postings to thousands of newsgroups and mailing lists. These attempts to grab free advertising are refered to as "spams" in the usual, somewhat boneheaded, net terminology. USENET is more susceptible to this practice, and many spams originate on the USENET groups and then are passed on to the mailing lists. However, spammers also get lists of mailing addresses and hit these too, so neither medium is immune. What should you do personally if you get junk mail? --------------------------------------------------- Just delete it and move on without reading it further. Filing a protest is becoming increasingly useless because spammers are often disguising the addresses where the messages are sent from. Unless you really understand Internet mail systems, your attempt at protest by sending replies to the message will often end up being sent to the address of an innocent person that the spammer is victimizing. What can BIOSCI/bionet do to protect its newsgroups? ---------------------------------------------------- The only solution currently available is to moderate the newsgroup. If this newsgroup is already moderated, then you are in good shape. Moderation protects the USENET distribution from about 95% of the spams that are being sent to date and protects the mailing lists completely. Moderation means, however, that someone has to take the time to review each message before it goes out. We have set up software here that simply allows the moderator to forward to an address at net.bio.net messages that (s)he wishes to have distributed. This takes no more time than that needed to read the message and pass it on, say about 1 min. per message. Most newsgroups currently have a discussion leader who is responsible for their newsgroup. The discussions leaders and their e-mail addresses are listed in the BIOSCI Information Sheet which is available on the Web at http://www.bio.net/. If a newsgroup is being hit with too many junk postings, please contact the discussion leader for that group and see if there is interest in moderating the group. Please do not assume that by simply posting a complaint to the newsgroup itself, anyone on the BIOSCI staff will act on your complaint. With close to 100 newsgroups to run, the BIOSCI staff has to rely on the discussion leaders of each newsgroup to report problems directly to us at biosci-help@net.bio.net. We will moderate any of our newsgroups if the discussion leader tells us that the readership of the group wishes to do so and if a moderator is willing to do the work. For most BIOSCI/bionet groups, this entails only a few minutes of work each day. Moderating a newsgroup will resolve probably 95% of the junk postings on the USENET distribution. Unfortunately there are easy ways for determined spammers to override the moderation mechanism on USENET, but we can protect our e-mail subscribers from unwanted postings if the newsgroup is moderated. You can also access our newsgroups over the WWW at URL http://www.bio.net. While this Web interface will not stop spammers from trying to post to the groups, this will give you yet another way, besides using USENET news, to keep the junk out of your personal mail files. For those of you with local USENET news systems, the Web interface will also give you faster access to new newsgroups and recent postings. 3) Examples of subscribing and unsubscribing to the mailing lists. ------------------------------------------------------------------ PLEASE NOTE: The BIOSCI management does NOT act on subscription/unsubscription requests that are posted improperly to the newsgroups and mailing lists. People who do this only bother everyone on the lists to no avail. Please be sure to follow the proper procedures below. Gory details are in the BIOSCI Information sheets on the Web at http://www.bio.net. Below we give an example utilizing the METHODS-AND-REAGENTS list at both of our two BIOSCI sites: Users in the Americas and Pacific Rim countries who use the BIOSCI ------------------------------------------------------------------ node at computer net.bio.net: ---------------------------- A) Determine the "listname" which is the <=8 character mail address ^^^^^^^^^^^^^ for the group. These can be found in the BIOSCI Info. Sheet. For the METHODS-AND-REAGENTS group the mailing address is methods@net.bio.net. The listname is the portion of the address to the left of the @ sign, i.e., "methods". The listname is used with the "subscribe" and "unsubscribe" commands illustrated below. B) Mail all commands in the body of a mail message addressed to biosci-server@net.bio.net. Do NOT send commands to the newsgroup posting addresses! Leave the Subject: line blank, any text on it will be ignored. C) In the body of your message put one or more of the following commands with an "end" command on the last line, e.g., subscribe methods unsubscribe methods end Do NOT put your e-mail address or other text on these lines. The server only allows you to cancel your subscription if the address on your mail header matches the address on our mailing list. Please ask for help at biosci-help@net.bio.net if your address has changed, e.g., if you know you are on the list but the server tells you that you are not a member. Users in Europe, Africa, and Central Asia who use the BIOSCI node at -------------------------------------------------------------------- computer daresbury.ac.uk (also known as dl.ac.uk): ------------------------------------------------- To subscribe and unsubscribe to/from the BIOSCI lists, you need to specify the full USENET newsgroup name with "bionet-news." prepended. The USENET newsgroup names are listed in the BIOSCI Information sheet on the Web at http://www.bio.net/. For the METHODS-AND-REAGENTS list the USENET newsgroup name is bionet.molbio.methds-reagnts, thus the appropriate commands are sub bionet-news.bionet.molbio.methds-reagnts unsub bionet-news.bionet.molbio.methds-reagnts These commands are included in a message addressed to mxt@dl.ac.uk, NOT to the newsgroup mailing addresses. As usual, include the text in the body of the message as text on the Subject: line is ignored. To unsubscribe from all the lists at the UK node, use unsub bionet-news Please note that if the address in the list is different than the one in your mail message header, you will not be able to unsubscribe by this method. If you have problems, please mail biosci@daresbury.ac.uk. 4) The BIOSCI user address and research interest directory. ----------------------------------------------------------- Please take this opportunity to add your name, address, and research interest information to the BIOSCI User Address Database if you have not already done so. You can fill out the address form directly through our Web page at URL http://www.bio.net/adrform.html. The address database is reindexed nightly for WWW access (the URL is http://www.bio.net/). If you are not directly on the Internet but can reach it by e-mail, please use our waismail server to access the user directory. waismail use is described above. You can also request a user address form by e-mail from biosci-help@net.bio.net. Please check your database entry from time-to-time to see if your address information is still up-to-date. Because of our limited personnel resources, we ask that you resubmit a *complete* form to revise your entry; we only replace complete entries and do not have resources to edit old forms. Sincerely, Dave Kristofferson BIOSCI/bionet Manager biosci-help@net.bio.net From owner-rapd@net.bio.net Thu Apr 17 23:00:00 1997 Path: biosci!AZUL.CTC.COM.BR!CARLOS.CTFT-1.CTFT.CTC From: CARLOS.CTFT-1.CTFT.CTC@AZUL.CTC.COM.BR Newsgroups: bionet.molbio.rapd Subject: AFLP adapters Date: 18 Apr 1997 04:24:02 -0700 Organization: Centro de Tecnologia Copersucar Lines: 18 Sender: daemon@net.bio.net Distribution: world Message-ID: <2393DC77856@azul.ctc.com.br> NNTP-Posting-Host: net.bio.net Would you know a company that synthesize adapters to perform the AFLP analysis of plant genome? Thank you. ================================ Carlos Gonzaga de Almeida, Ph.D. Plant Science Copersucar Technology Center P.O.Box 162 Piracicaba, SP 13400-970 BRAZIL ================================ phone:+55 194 298212 fax :+55 194 298388 email: gonzaga@azul.ctc.com.br ================================ From owner-rapd@net.bio.net Thu Apr 17 23:00:00 1997 Path: biosci!CGNET.COM!J.PHAM From: J.PHAM@CGNET.COM (Jean-Louis Pham) Newsgroups: bionet.molbio.rapd Subject: AFLP in routine Date: 17 Apr 1997 19:26:24 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 15 Sender: daemon@net.bio.net Distribution: world Message-ID: <01BC4BE3.405C9540@GRC.irri.cgiar.org> NNTP-Posting-Host: net.bio.net Would you know a company that could perform the AFLP analysis of a few hundreds of samples ? Thank you. Jean-Louis Pham ---------------------------------------------------------------------------------------------------------- Genetic Resources Center Fax: (63-2) 891-1292 International Rice Research Institute (63-2) 845-0606 P.O. Box 933 (63-2) 761-2406 1099 Manila Philippines E-mail: j.pham@cgnet.com ----------------------------------------------------------------------------------------------------------- From owner-rapd@net.bio.net Fri Apr 18 23:00:00 1997 Path: biosci!cpsarg.com!manuel&lilianacriado From: manuel&lilianacriado@cpsarg.com ("Liliana Martinez y Manuel Criado") Newsgroups: bionet.molbio.rapd Subject: about RAPD in onion Date: 19 Apr 1997 14:43:06 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 14 Sender: daemon@net.bio.net Distribution: world Message-ID: <19970419215032480.AAA170@ppp-16.mza.satlink.com> NNTP-Posting-Host: net.bio.net I am working with RAPD´s molecular markers, obtained by PCR, linked to pink root resistant in onion. I have done many reactions of PCR but the yield of the band in the electroforesis are not strong. The amplified reactions function, so I have a lot of polimorfics bands, but they are so weak. I have probed differents Taq polimerase, nucleotides, buffers, water qualities, primers and DNA with and without cleaning, but the bands are still weaks. I do not what is the reason of this result. Do you have any experience with this problem. Looking forward to hearing from you, Liliana From owner-rapd@net.bio.net Sun Apr 20 23:00:00 1997 Path: biosci!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!feed1.news.erols.com!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!newsxfer3.itd.umich.edu!portc01.blue.aol.com!audrey02.news.aol.com!not-for-mail From: punkinn97@aol.com (Punkinn97) Newsgroups: bionet.molbio.rapd Subject: RAPD techniques Date: 21 Apr 1997 03:33:51 GMT Organization: AOL http://www.aol.com Lines: 11 Message-ID: <19970421033200.XAA19570@ladder01.news.aol.com> NNTP-Posting-Host: ladder01.news.aol.com X-Admin: news@aol.com I am currently using RAPD in analyzing bird DNA. Has anyone else used RAPD for these purposes? Also, I am trying to get a feel for how reliable people think RAPD is. I am a senior in college and I am having a hard time finding information re: RAPDs reliability. Also, we are having a hard time getting bands and I am interested in seeing if other people, using Mammalian DNA , are having any luck. If you can answer any of these questions, please right back. Thank you , The Forever Frustrated Biology Major Kristin Lange From owner-rapd@net.bio.net Sun Apr 20 23:00:00 1997 Path: biosci!daresbury!uninett.no!news-feed.inet.tele.dk!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!news-peer.gsl.net!news-hk.gsl.net!news.gsl.net!newsgate.cuhk.edu.hk!news.cuhk.edu.hk!usenet From: He Xian Hui Newsgroups: bionet.molbio.rapd Subject: looking for good protocol(s) for isolation of DNA from dry medicinal plants Date: Mon, 21 Apr 1997 22:21:42 -0700 Organization: Department of Physiology, Chinese University of Hong Kong, Shatin, N. T., HONG KONG Lines: 12 Message-ID: <335C4AE6.60D3@cuhk.edu.hk> Reply-To: hexianhui@cuhk.edu.hk NNTP-Posting-Host: @mt_lab2.psi.cuhk.edu.hk Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 3.01Gold (Win95; I; 16bit) Dear Netters, When isolating genomic DNA from dry medicinal plants using SDS or CTAB method, I fund that the total DNA was always sheared heavily. Sometimes the pattern of the electrophoresis of isolated DNA did not show any bands other than smearing.I don't why. Would you please send some good ideas or protocol(S)related to this question? Thanks a lot. Rong-Zhao Fu email: z044602@cuhk.edu.hk From owner-rapd@net.bio.net Sun Apr 20 23:00:00 1997 Path: biosci!ihnp4.ucsd.edu!munnari.OZ.AU!news.ecn.uoknor.edu!feed1.news.erols.com!news.maxwell.syr.edu!ott.istar!news.istar.net!tor.istar!east.istar!news1.istar.ca!news From: Frank BERAUD Newsgroups: bionet.molbio.rapd Subject: Re: about RAPD in onion Date: 20 Apr 1997 17:47:10 GMT Organization: iSTAR Internet Incorporated Lines: 21 Message-ID: <5jdkqu$2dd@news.istar.ca> References: <19970419215032480.AAA170@ppp-16.mza.satlink.com> Reply-To: pebio5@istar.ca NNTP-Posting-Host: ts4-12.mtl.istar.ca Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Mailer: Mozilla 3.0Gold (Win95; I) Liliana Martinez y Manuel Criado wrote: > > I am working with RAPD´s molecular markers, obtained by PCR, linked to pink > root resistant in onion. > I have done many reactions of PCR but the yield of the band in the > electroforesis are not strong. The amplified reactions function, so I have > a lot of polimorfics bands, but they are so weak. > I have probed differents Taq polimerase, nucleotides, buffers, water > qualities, primers and DNA with and without cleaning, but the bands are > still weaks. > I do not what is the reason of this result. > Do you have any experience with this problem. > > Looking forward to hearing from you, > > Liliana Did you try the AmpiTaq Stoffel Fragment (Perkin Elmer) which lacks the 5' exo activity and gives you sharper bands and better discrimination ? It works pretty well. Frank From owner-rapd@net.bio.net Mon Apr 21 23:00:00 1997 Path: biosci!ACD.TUSK.EDU!prakash From: prakash@ACD.TUSK.EDU ("C. S. Prakash") Newsgroups: bionet.molbio.rapd Subject: A New Newsgroup for Biotech in India Date: 22 Apr 1997 16:37:04 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 37 Sender: daemon@net.bio.net Distribution: world Message-ID: NNTP-Posting-Host: net.bio.net Dear Colleagues: A new internet discussion group to promote biotechnology in India has been started by Prof. DPS Verma of Ohio State University. To goal is to stimulate interaction and discussion on various opportunities and constraints towards the development of Agricultural Biotechnology in India. To subscribe to this list (called PBASIO--"Plant Biotechnologists and Agricultural Scientists of Indian Origin") send an e-mail to: listserver@lists.acs.ohio-state.edu Type the following in the body of the message text (do not give any subject heading): subscribe PBASIO Please feel free to pass this message around. Yours, C. S. Prakash Tuskegee University prakash@acd.tusk.edu --------------------------------------------- C. S. Prakash Prakash@Acd.Tusk.Edu Center for Plant Biotechnology Research Tuskegee University Phone (334) 727 8023 College of Agriculture Fax (334) 727 8552 Tuskegee, AL 36088. USA http://agriculture.tusk.edu ---------------------------------------------- From owner-rapd@net.bio.net Mon Apr 21 23:00:00 1997 Path: biosci!bcm.tmc.edu!cs.utexas.edu!news.maxwell.syr.edu!hunter.premier.net!uunet!in1.uu.net!152.163.170.17!newstf01.news.aol.com!audrey02.news.aol.com!not-for-mail From: punkinn97@aol.com (Punkinn97) Newsgroups: bionet.molbio.rapd Subject: Reliablity of RAPD Date: 22 Apr 1997 01:17:25 GMT Organization: AOL http://www.aol.com Lines: 10 Message-ID: <19970422011700.VAA29625@ladder01.news.aol.com> NNTP-Posting-Host: ladder01.news.aol.com X-Admin: news@aol.com I guess my real question concerning RAPDs reliability stems from the fact that it is a rather recent technology. From what I can tell, it does appear to be reliable in most circumstances, if all of the reagents are at the right concentrations. Does anyone share that opinion? Or has anyone heard that the slower, heated block or water bath systems tend to give better more reliable results? Thank you so much for all your help! It is GREATLY appreciated! Take care, Kristin From owner-rapd@net.bio.net Tue Apr 22 23:00:00 1997 Path: biosci!AGERI.SCI.EG!idl From: idl@AGERI.SCI.EG (Abdel Mohsen Tohamy) Newsgroups: bionet.molbio.rapd Subject: Reliability of RAPD-PCR Date: 23 Apr 1997 02:02:09 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 21 Sender: daemon@net.bio.net Distribution: world Message-ID: <199704230722.JAA15918@ageri.sci.eg> NNTP-Posting-Host: net.bio.net Dear Christin and all netters: Hello. In fact your question about the reliability of RAPD-PCR was one of my questions to the RAPD discussion group. This question had had many replies fron the list and I am looking for these replies to send it to you. Generally, the most of opinions had informed me that RAPD can be used to study the diversity between the organism of different taxa and for drawing the phylogenitic tree but RFLP is more reliable in case of studying the Phylogeny. Unfortunately, I have lost your Email. So, Please, send me your Email address again to send you these replies in case of finding it. With my best wishes. Nasser ************************************************************* Abd El-Nasser M. Elashry Agricultural Genetic Engineering Research Institute (AGERI) Agricultural Research Center (ARC) 9 Gamaa st., Giza 12619 Egypt Email: idl@ageri.sci.eg ************************************************************* From owner-rapd@net.bio.net Tue Apr 22 23:00:00 1997 Path: biosci!bcm.tmc.edu!cs.utexas.edu!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!feed1.news.erols.com!news.idt.net!usenet.logical.net!news.dal.ca!newsflash.concordia.ca!SUNqbc.risq.net!news.risq.qc.ca!news.mcgill.ca!feed.umontreal.ca!alize.ERE.UMontreal.CA!landryp From: landryp@ERE.UMontreal.CA (Landry Pierre-Alexandre) Newsgroups: bionet.molbio.rapd Subject: Re: Reliablity of RAPD Date: 23 Apr 97 14:53:42 GMT Organization: Universite de Montreal Lines: 28 Distribution: world Message-ID: References: <19970422011700.VAA29625@ladder01.news.aol.com> NNTP-Posting-Host: alize.ere.umontreal.ca punkinn97@aol.com (Punkinn97) writes: >I guess my real question concerning RAPDs reliability stems from the fact >that it is a rather recent technology. From what I can tell, it does >appear to be reliable in most circumstances, if all of the reagents are at >the right concentrations. Does anyone share that opinion? Or has anyone >heard that the slower, heated block or water bath systems tend to give >better more reliable results? >Thank you so much for all your help! >It is GREATLY appreciated! I do not think that questions arising from RAPDs are related to the age of this technique: numerous studies published in Biotechniques have explored almost all the possible parameters of the reaction that can be modified. The conclusion is always the same: you need to optimize your reaction to get reliable results. But I still think that the most important factor is the quality and the quantity of the template DNA your are amplifying (although none of these studies have discussed the importance to wear the powerful RAPD amulet ;) Pierre-Alexandre Landry Dept. de Sciences Biologiques Universite de Montreal C.P. 6128, succursale centre ville Montreal, Quebec H3C 3J7 PCR is just modern wizardry From owner-rapd@net.bio.net Tue Apr 22 23:00:00 1997 Path: biosci!rutgers.rutgers.edu!gatech!csulb.edu!hammer.uoregon.edu!zephyr.texoma.net!news.spinne.com!news.maxwell.syr.edu!news.radio.cz!CESspool!news.uakom.sk!classic.fris.sk!krajmer From: Diana Krajmerova Newsgroups: bionet.molbio.rapd Subject: thermal homogeneity Date: Wed, 23 Apr 1997 09:03:25 +0200 Organization: UAKOM Banska Bystrica Lines: 18 Message-ID: References: <3.0.32.19970131094309.0068c658@ucspop.byu.edu> <32F2DDA6.740F@unity.ncsu.edu> NNTP-Posting-Host: classic.fris.sk Mime-Version: 1.0 Content-Type: TEXT/PLAIN; charset=US-ASCII In-Reply-To: <32F2DDA6.740F@unity.ncsu.edu> We just plan to buy a cykler for the fist time, we do not have any experience. Would anybody tell me if thermal homogeneity across the block +-0.5 C is O.K.? So far we plan to do RAPDs for genotype checks in seed orchards and somaclonal variability. Thanks. Diana ---------------------------------------------------------------- Diana KRAJMEROVA Tel.: +42 855 314 333 Forest Research Institute Fax.: +42 855 321 883 T.G.Masaryka 22 960 92 Zvolen Slovakia e-mail: krajmer@fris.sk From owner-rapd@net.bio.net Wed Apr 23 23:00:00 1997 Path: biosci!daresbury!uninett.no!news-feed.inet.tele.dk!cpk-news-hub1.bbnplanet.com!news.bbnplanet.com!news-peer.sprintlink.net!news.sprintlink.net!sprint!howland.erols.net!newsfeed.internetmci.com!news.cyberhighway.net!not-for-mail From: Robert Clark Newsgroups: bionet.molbio.rapd Subject: Re: about RAPD in onion Date: Tue, 22 Apr 1997 14:12:01 -0700 Organization: CyberHighway Internet Services Lines: 63 Message-ID: <335D29A1.16A7@cyberhighway.net> References: <19970419215032480.AAA170@ppp-16.mza.satlink.com> NNTP-Posting-Host: 206.26.230.7 Mime-Version: 1.0 Content-Type: text/plain; charset=iso-8859-1 Content-Transfer-Encoding: 8bit X-Mailer: Mozilla 3.0 (Win16; I) To: Liliana Martinez y Manuel Criado Liliana Martinez y Manuel Criado wrote: > > I am working with RAPD´s molecular markers, obtained by PCR, linked to pink > root resistant in onion. > I have done many reactions of PCR but the yield of the band in the > electroforesis are not strong. The amplified reactions function, so I have > a lot of polimorfics bands, but they are so weak. > I have probed differents Taq polimerase, nucleotides, buffers, water > qualities, primers and DNA with and without cleaning, but the bands are > still weaks. > I do not what is the reason of this result. > Do you have any experience with this problem. > > Looking forward to hearing from you, > > Liliana Hello Liliana: There are many possibilities. Here are some aspects to consider. You have already mentioned some of them, but I list them because they were our trouble points when we experienced similar problems. Another point to consider is that the onion genome is HUGE. I'm not sure what this would do to the amount of each reaction product. We do not do onion RAPDs in our lab; the following notes hold true for other veg. species. 1. Concentration of your onion DNA. We try to get 25 ng of template in a 25 ul total reaction volume. 2. Purity of your onion DNA. How are you getting the DNA? From what tissue? We have found that for RAPDs especially, to get the best band intensity you must use DNA that is very clean. By this, I mean without visible pigments or debris in the pellet. We often use a CTAB procedure for larger samples (1-2 grams leaf tissue), but we have also found that an SDS-NaCl-EDTA-isopropanol procedure works well with smaller samples. We have also found that one single DNA prep procedure does not work across different plant species, probably because different species contain different PCR inhibitors. 3. Type of polymerase. We routinely use Stoffel fragment instead of the complete Taq polymerase. The processivity is slower, but this truncated enzyme tolerates higher melt temperatures and higher Mg concentrations (see below). 4. Magnesium concentration. 4-5 mM with Stoffel cured our repeatability and band intensity problems for most RAPD reactions. 5. Thermal cycle optimization. Not all RAPD primers respond in a cooperative way to the same temperature cycle. You may have to experiment to find the ideal. We found a general pattern is to start with a relatively low annealing temperature (40 C) for the first 5 cycles, then increase the annealing temperature stringency for the remaining cycles (50 C). 6. Type of stain for your gel, and type of gel. We have found that the combination of polyacrylamide gels and silver staining is vastly superior to agarose and ethidium bromide. The sensitivity using the PAGE-silver staining system I think must be 10 to 20 times greater than the other system. Good luck! --Bob From owner-rapd@net.bio.net Wed Apr 23 23:00:00 1997 Path: biosci!daresbury!bioftp.unibas.ch!ubaclu.unibas.ch!locherr From: locherr@ubaclu.unibas.ch (NAME) Newsgroups: bionet.molbio.rapd Subject: software request Message-ID: <1997Apr24.181135.47483@yogi.urz.unibas.ch> Date: 24 Apr 97 18:11:35 MET Organization: University of Basel, Switzerland Lines: 17 Hi out there in the scientific world In our lab in Basel, one of the main research field consists on progeny array analysis and on multiple paternity determination. Our analysis are realised on mother and their sibling (putative fathers unknown). Our genetic data are inferred from dominant markers (RAPDs). Our questions are : Does a software exist dealing with such kind of data and answering the following question: -How many fathers are there ? and -Confidence limits ? Thanks for any kind of help. Rolf Locher + Thierry Wirth