From owner-biotechniques@net.bio.net Wed Aug 13 23:00:00 1997 Path: biosci!biosci!not-for-mail From: Anne Siri Langelandsvik Newsgroups: bionet.journals.letters.biotechniques Subject: Automated fluorescent primer extension Date: 14 Aug 1997 05:39:19 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 23 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <9708140756.AA25351@nlh10.nlh.no> NNTP-Posting-Host: net.bio.net Hi, I am going to do an automated fluorescent primer extension, that is using fluorescent labeled primer for the primer extension so that I can detect the band using an automatic DNA sequencer. However,do I need to use fluor. labeled primer for the DNA sequencing aswell,or can I do the "one" tube sequencing reaction involving fluor. labeled ddNTP's? The reason for this question is that I wonder if the extension product versus the sequencing products will run different because the primer is labeled with a "big" fluorochrom.=20 Anne Siri ................................................................... Anne Siri Langelandsvik Laboratory of Microbial Gene Technology Agricultural University of Norway P.O. Box 5051, N-1432 =C5s Tel: +47 64 94 85 96/94 69 Fax:: +47 64 94 14 65 Homepage: http://wwwnlh.nlh.no/institutt/ibf/ansatte/AL/ibf_ala.htm Private: S=F8r=E5sveien 18, N-1430 =C5s Tel: +47 64 94 45 16 ................................................................... From owner-biotechniques@net.bio.net Thu Aug 14 23:00:00 1997 Path: biosci!biosci!not-for-mail From: MMCCARTHY@biotechnet.com Newsgroups: bionet.journals.letters.biotechniques Subject: BioTechniques 23(3), September 1997 Date: 15 Aug 1997 05:25:10 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 191 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <01IMFN1JNWF68ZHC2W@delphi.com> NNTP-Posting-Host: net.bio.net BioTechniques 23(3), September 1997 Table of Contents Benchmarks Automated Fluorescence-Based Screening for Mutation by SSCP: Use of Universal M13 Dye Primers for Labeling and Detection P. Boutin, E.H. Hani, F. Vasseur, C. Roche, B. Bailleul, J. Hager and P. Froguel DNA Sequencing with Modular Primers Using a Two-Step Protocol with Thermostable Polymerase at the Second Step M.C. Raja, D. Zevin-Sonkin, J. Shvartzburd, L. Kotler and L. Ulanovsky Sequencing of 3-prime cDNA Clones Using Anchored Oligo(dT) Primers J. Liao and Z. Gong Post-PCR Labeling Using Taq DNA Polymerase Z. Chen and D.E. Ruffner Identification of Targeted Embryonic Stem Cells Using Long-Distance PCR E.V. Loukianov, T. Loukianova and M. Periasamy Enhancement of PCR-RFLP Typing of Bovine Leukocyte Adhesion Deficiency A. Zsolnai and L. Fesus Directional Immobilization of Heparin onto the Nonporous Surface of Polystyrene Microplates V.D. Nadkarni and R.J. Linhardt Enhancement of Diaminobenzidine Colorimetric Signal in Immunoblotting L.A. Pukac, J.E. Carter, K.S. Morrison and M.J. Karnovsky Even Highly Homologous Proteins Vary in their Affinity for Different Blotting Membranes N. Balasubramanian and S.M. Zingde Useful Vectors for the Two-Hybrid System in Mammalian Cells M. Buchert, S. Schneider, M.T. Adams, H.P. Hefti, K. Moelling and C.M. Hovens Simultaneous Expression of Multi-Subunit Proteins in Mammalian Cells Using a Convenient Set of Mammalian Cell Expression Vectors M.I. Cockett, R. Ochalski, K. Benwell, R. Franco and J. Wardwell-Swanson Site-Directed Mutagenesis: A Two-Step Method Using PCR and DpnI S. Li and M.F. Wilkinson PCR-Based Random Mutagenesis Using Manganese and Reduced dNTP Concentration J.L. Lin-Goerke, D.J. Robbins and J.D. Burczak Near-Zero Background Cloning of PCR Products Q. Zeng, M.K. Eidsness and A.O. Summers Ligation of 5-prime DNA Overhangs to 3-prime DNA Overhangs Using Short Single-Stranded Oligonucleotide Linkers G. Zhao and M. Hortsch Nuclei Isolation from Bone Cells for Nuclear Run-on Assays N. Elango, J. Vivekananda, R. Strong and M.S. Katz Ammonium Acetate Protocol for the Preparation of Plasmid DNA Suitable for Mammalian Cell Transfections S.M. Saporito-Irwin, R.T. Geist and D.H. Gutmann Rapid Purification of Reporter Group-Tagged Inositol Hexakisphosphate on Ion-Exchange Membrane Adsorbers A. Chaudhary, B. Mehrotra and G.D. Prestwich Modified Semliki Forest Virus Expression Vector that Facilitates Cloning J. Meanger, I. Peroulis and J. Mills Set of Optimized Luciferase Reporter Gene Plasmids Compatible with Widely Used CAT Vectors J. Altschmied and J. Duschl The Internet On-Ramp Cyberspace for Biologists Short Technical Reports Quantification of Transcript-to-Template Ratios as a Measure of Gene Expression Using RT-PCR I. Harting and R.J. Wiesner Design and Testing of beta-Actin Primers for RT-PCR that Do Not Co-amplify Processed Pseudogenes T. Raff, M. van der Giet, D. Endemann, T. Wiederholt and M. Paul Use of Two Reverse Transcriptases Eliminates False-Positive Results in Differential Display Y.-J. Sung and R.B. Denman High-Stringency Subtraction for the Identification of Differentially Regulated cDNA Clones C.P. Scutt and P.M. Gilmartin Measurement of Telomeric DNA Content in Human Tissues J.E. Bryant, K.G. Hutchings, R.K. Moyzis and J.K. Griffith Controlled Gene Gun Delivery and Expression of DNA Within the Cornea D.L. Tanelian, M.A. Barry, S.A. Johnston, T. Le and G. Smith Phosphorylation-Directed Antibodies in High-Flux Screens for Compounds that Modulate Signal Transduction J.A. Alberta and C.D. Stiles One Primer Pair Amplifies Small Subunit Ribosomal DNA from Mitochondria, Plastids and Bacteria P. Berschick Analysis of Assembly of Synthetic Antibody Fragments: Expression of Functional scFv with Predefined Specificity N. Kobayashi, E. Soderlind and C.A.K. Borrebaeck Research Reports Multiplex PCR: Critical Parameters and Step-by-Step Protocol O. Henegariu, N.A. Heerema, S.R. Dlouhy, G.H. Vance and P.H. Vogt PNA-Mediated Purification of PCR Amplifiable Human Genomic DNA from Whole Blood C. Seeger, H.-G. Batz and H. Orum In-Cell Assembly of scFv from Human Thyroid Infiltrating B Cells N. Chapal, M. Bouanani, M.J. Embleton, I. Navarro-Teulon, M. Biard-Piechaczyk, B. Pau and S. Peraldi-Roux Product Application Focus Early Detection of Apoptosis Using a Fluorescent Conjugate of Annexin V G. Zhang, V. Gurtu, S.R. Kain and G. Yan Fluorescent Dye Assay for Detection of DNA in Recombinant Protein Products R. Bolger, F. Lenoch, E. Allen, B. Meiklejohn and T. Burke New Products Index to Advertisers From owner-biotechniques@net.bio.net Tue Aug 19 23:00:00 1997 Path: biosci!biosci!not-for-mail From: BIOSCI Administrator Newsgroups: bionet.journals.letters.biotechniques Subject: BIOSCI/bionet miniFAQ & Fundraiser Date: 20 Aug 1997 06:54:30 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 233 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <199708200900.CAA12390@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.