From owner-biotechniques@net.bio.net Wed May 07 23:00:00 1997 Path: biosci!biosci!not-for-mail From: wfischer@sunflower.bio.indiana.edu (Will Fischer) Newsgroups: bionet.journals.letters.biotechniques Subject: Re: protoplast buffer and lysis buffer Date: 8 May 1997 06:13:12 -0700 Organization: Biology, Indiana University - Bloomington Lines: 44 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <5kqij5$euh$1@dismay.ucs.indiana.edu> References: <3365E158.5F4A@gems.vcu.edu> NNTP-Posting-Host: net.bio.net Robyn Puffenbarger (rpuffenbarg@Gems.VCU.EDU) wrote: : : I have the protocol for the "in-well-lysis" procedure, but have lost the : recipies for the protoplast buffer and the lysis buffer. This simple : technique lets you quickly see the difference in recombinant plasmids : without a 'miniprep.' Protoplasting buffer: 30 mM Tris-HCl pH 8.0 5 mM Na2EDTA 50 mM NaCl 20 % sucrose 50 ug/ml RNase A 50 ug/ml lysozyme (I also add 1 mg/ml Orange G) Lysis buffer: 89 mM Tris 89 mM boric acid 2.5 mM Na2EDTA 2 % SDS 5 % sucrose 0.04 % bromophenol blue Reference: Sekar, V. (1987) BioTechniques 5:11-13 Alternative technique: Pick colonies for PCR with vector primers (protocol available at http://www.bio.indiana.edu/~wfischer/protocol_colony-PCR-screen.html). This allows preparation of sequencing template as part of the screening step! -- ____________________________________________________________ Will Fischer Biology Department wfischer@indiana.edu Jordan Hall http://www.bio.indiana.edu/~wfischer Indiana University Lab: 812-855-2549 Bloomington, Indiana 47405 USA FAX: 812-855-6705 From owner-biotechniques@net.bio.net Fri May 16 23:00:00 1997 Path: biosci!biosci!not-for-mail From: MMCCARTHY@biotechnet.com Newsgroups: bionet.journals.letters.biotechniques Subject: BioTechniques 22(6), June 1997 Date: 16 May 1997 18:19:28 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 185 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <01IIY9O9IE2Q9ELIVL@delphi.com> NNTP-Posting-Host: net.bio.net BioTechniques 22(6), June 1997 Table of Contents BENCHMARKS In situ Trypan Blue staining of monolayer cell cultures for permanent fixation and mounting Perry, S.W., Epstein, L.G., and Gelbard, H.A. Detection of specific DNA-binding protein in HeLa whole cell extract Gai, Q., Xu, G., Shannon, T., and James, C.B.L. Optimized conditions of PacI and SwaI for genomic analysis of X. axonopodis pv. vesicatoria by PFGE Hacioglu, E., Basim, H., and Stall, R.E. Screening of yeast transformants by chemiluminescence for detection of secreted heterologous proteins Arffman, A., Uitto, J., and Aho, S. Improved cloning vectors for transgene construction Yang, Y. and Spector, A. PCR strategy to obtain the 5' and 3' flanking sequences of partial clones from lambdaZAP cDNA libraries Li, Q. and Nicholas, R.A. Partial inverse PCR: a technique for cloning flanking sequences Pang, K.M. and Knecht, D.A. Refinements in re-amplification and cloning of DDRT-PCR products Bhattacharjee, A., Rutherford, M.S., Abrahamsen, M.S., Lappi, V.R., and Schook, L.B. PCR mutagenesis: treatment of the megaprimer with mung bean nuclease improves yield Chattopadhyay, D., Raha, T., and Chattopadhyay, D. Production of PCR mimics for any semiquantitative PCR application Ali, S.A., Sarto, I., and Steinkasserer, A. Effects of gelatin and BSA on the amplification reaction for generating RAPD Stommel, J.R., Panta, G.R., Levi, A., and Rowland, L.J. Determination of fibroblast growth factor receptor expression in mouse, rat and human samples using a single primer pair McEwen, D.G. and Ornitz, D.M. Analysis of primer-derived nonspecific amplification products in RAPD-PCR Pan, Y.-B., Burner, D.M., Ehrlich, K.C., Grisham, M.P., and Wei, Q. Isolation of thermophilic bacteria using bacteriological grade agar at temperatures above 80=F8C D'Souza, D.R., Kaur, R., Pajni, S., and Sharma, P. Extraction of DNA using monoclonal anti-DNA and magnetic beads Gelsthorpe, A.R., Gelsthorpe, K., and Sokol, R.J. Rapid isolation of total RNA from small samples of hypocellular, dense connective tissue Reno, C., Marchuk, L., Sciore, P., Frank, C.B., and Hart, D.A. Evaluation of DNA sequencing ambiguities using tetramethylammonium chloride hybridization conditions Connors, T.D., Burn, T.C., VanRaay, T., Germino, G.G., Klinger, K.W., and Landes, G.M. THE INTERNET ON-RAMP Cyberspace for Biologists SHORT TECHNICAL REPORTS Simultaneous in situ detection of apoptosis and necrosis in monolayer cultures by TUNEL and Trypan Blue staining Perry, S.W., Epstein, L.G., and Gelbard, H.A. High-throughput RT-PCR analysis of multiple transcripts using a microplate RNA isolation procedure Su, S., Vivier, R.G., Dickson, M.C., Thomas, N., Kendrick, M.K., Williamson, N.M., Anson, J.G., Houston, J.G., and Craig, F.F. Isolation of high molecular weight DNA for genotyping of transgenic mice Malumbres, M., Mangues, R., Ferrer, N., Lu, S., and Pellicer, A. Analysis of the size distribution of first-strand cDNA molecules Bodescot, M. and Brison, O. Use of manganese in RT-PCR eliminates PCR artifacts resulting from DNaseI digestion Bauer, P., Rolfs, A., Regitz-Zagrosek, V., Hildebrandt, A., and Fleck, E. RESEARCH REPORTS Cassette for the generation of sequential gene disruptions in the yeast Schizosaccharomyces pombe McNabb, D.S., Pak, S., and Guarente, L. Double-labeled fluorescent probes for 5' nuclease assays: purification and performance evaluation Rudert, W.A., Braun, E.R., Faas, S.J., Menon, R., Jaquins-Gerstl, A., and Trucco, M. BIOCOMPUTING Computer-assisted, quantitative cytokine enzyme-linked immunospot analysis of human immune effector cell function Cui, Y. and Chang, L.-J. Control of a remote microscope over the Internet Maturo, R., Kath, G., Zeigler, R., and Meechan, P. Computer program for calculating the melting temperature of degenerate oligonucleotides used in PCR or hybridization Chen, H. and Zhu, G. Rescuing corrupted gel files from Model 377 and 373 DNA sequencers O'Brien, K.M., Fondon III, J.W., Garner, H.R., and Evans, G.A. Visual Genome ExplorerTM, a comparative visual interface to genome data Archer, E.S., Yee, L.W., and Leung, F.C. PRODUCT APPLICATION FOCUS Application of oligonucleotide activation to restriction endonuclease NarI Senesac, J.H. and Romanin, J.K. Measurement of nucleic acid concentrations using the DyNAQuant(TM) and the GeneQuant(TM) Teare, J.M., Islam, R., Flanagan, R., Gallagher, S., Davies, M.G., and Grabau, C. 1997 AUTHORS INDEX (Volume 22) NEW PRODUCTS INDEX TO ADVERTISERS From owner-biotechniques@net.bio.net Mon May 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 May 1997 06:46:30 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 239 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <199705200900.CAA25885@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-biotechniques@net.bio.net Wed May 21 23:00:00 1997 Path: biosci!biosci!not-for-mail From: David Kristofferson Newsgroups: bionet.journals.letters.biotechniques Subject: IMPORTANT - BIOSCI moving to Stanford! Date: 22 May 1997 05:58:57 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 55 Sender: daemon@net.bio.net Approved: MMCCARTHY@BIOTECHNET.COM Distribution: world Message-ID: <199705220406.VAA04616@net.bio.net> NNTP-Posting-Host: net.bio.net After more than a decade of serving the biology community on the BIOSCI project, I have decided that it is time for me to pass the torch. The demands of my career and family are making it increasingly difficult for me to run BIOSCI adequately on a part-time basis, and I've decided that I would do the project more harm than good by clinging to it. Therefore I have concluded an agreement with the Stanford University Libraries to take over the management of the BIOSCI project effective 6 June 1997. Many of you know the work of the Stanford Libraries through HighWire Press which has done an excellent job bringing the Journal of Biological Chemistry, Science Magazine, and other prestigious journals to the WWW (see http://highwire.stanford.edu/). I have every confidence that Stanford will provide an environment where the BIOSCI project can flourish. The BIOSCI team at Daresbury will continue to support the project in Europe, and Dave Mack and Julie Lawrence on the current BIOSCI staff will continue to assist the new team at Stanford. The project will be managed by Serge Taylor of the Stanford Libraries who is responsible for developing Web-based "knowledge environments" for scientists. I will still participate with the project in an advisory capacity but have to phase myself out of day-to-day operations. Many of you who read bionet.announce, bionet.biology.computational, and bionet.jobs.offered (the groups that I moderate) have been aware of the recent posting delays that my busy schedule has inflicted on those newsgroups. I am pleased to announce that Todd McGee, Scientific Advisor of the HighWire Press, will be taking over moderation of those groups from me this week. ** NOTE ** - The BIOSCI hardware will be down for backups on 6 June and will be moved over the weekend of June 7 and 8. We will be changing the IP number of the machine, but the host name and all mailing addresses will remain unchanged. There may be a few days of service disruption during this time due to the move unfortunately, just as we experienced in BIOSCI's previous move. We hope to keep this to a minimum. We will post status reports on the move to bionet.announce before and after it happens. Please watch that newsgroup for details. It has been a pleasure serving all of you since the "early days" of biology on the Internet. I've had the pleasure of not only "knowing" many of you over the Internet, but have also met literally thousands of biologists around the U.S. and elsewhere during the many Internet training seminars that I gave over the last decade. I've been truly fortunate to have been given the chance to play a role in the growth of biology on the Internet. This is something I will always treasure. Sincerely, Dave Kristofferson BIOSCI/bionet Manager biosci-help@net.bio.net