From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Path: biosci!daresbury!yama.mcc.ac.uk!bofh.dot!thor.cf.ac.uk!news From: Nick Jacobsen Subject: Re: DNA ligation help Sender: news@cf.ac.uk (USENET News System) Message-ID: Date: Mon, 3 Jun 1996 09:21:04 GMT To: xcl@med.unc.edu X-Nntp-Posting-Host: d053.mg.cf.ac.uk Content-Transfer-Encoding: 7bit Content-Type: text/plain; charset=us-ascii References: <4opneu$nke@newz.oit.unc.edu> Mime-Version: 1.0 X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Organization: Institute of Medical Genetics Cardiff Lines: 14 Xref: biosci bionet.molbio.ageing:2744 bionet.molbio.bio-matrix:732 bionet.molbio.evolution:4576 bionet.molbio.gene-linkage:1080 bionet.molbio.methds-reagnts:45236 Hi, I find that the majority of problems with DNA ligation inefficiency are due to ATP degradation in the reaction buffer. If in doubt order new buffer and freeze in small aliquots. You can also order Pharmacia ligase - you have to order separate 10mM ATP with it to add to the reaction as required. You have to be fairly accurate with the amount of ATP added as too much is as bad as too little. I hope this is of help to you cheers Nick Jacobsen. From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Path: biosci!bcm.tmc.edu!pendragon!news.msfc.nasa.gov!elroy.jpl.nasa.gov!lll-winken.llnl.gov!fnnews.fnal.gov!unixhub!news.Stanford.EDU!not-for-mail From: ladasky@leland.Stanford.EDU (John Ladasky) Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Subject: Re: DNA ligation help Followup-To: bionet.molbio.methds-reagnts Date: 3 Jun 1996 10:47:29 -0700 Organization: Stanford University, CA 94305, USA Lines: 23 Message-ID: <4ov8fh$7gm@elaine35.Stanford.EDU> References: <4opneu$nke@newz.oit.unc.edu> NNTP-Posting-Host: elaine35.stanford.edu Xref: biosci bionet.molbio.ageing:2745 bionet.molbio.bio-matrix:733 bionet.molbio.evolution:4577 bionet.molbio.gene-linkage:1081 bionet.molbio.methds-reagnts:45260 Followups have been limited to bionet.molbio.methds-reagnts. In article <4opneu$nke@newz.oit.unc.edu>, chunlin xin wrote: >Hi, there, I have trouble in DNA ligation, the case below: > Insert Fragment: 16.3kb SalI Frag. > Vector: Bscrip-KS(-) SalI cut plasmid or other > salI cut expression vector > It looks like no ligation problem, but in fact, I cannot >success to ligate them and transfer them into DH5a no matter I use >chemical or electroparation method, Have anybody ever meet the same problem? >How can I solve them? Any suggestion is appreciated! Thanks for you attention! A 16.3 Kb insertion is quite large. DH5-alpha, and most other normal strains of E. coli, cannot reliably maintain plasmids larger than 10 Kb. (I'm not sure why this is so.) So your ligation and transformation may both be successful, but then you are not getting any ampicillin-resistant colonies. Can you subclone this fragment? -- Unique ID : Ladasky, John Joseph Jr. Title : BA Biochemistry, U.C. Berkeley, 1989 (Ph.D. perhaps 1998???) Location : Stanford University, Dept. of Structural Biology, Fairchild D-105 Keywords : immunology, music, running, Green From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Path: biosci!daresbury!yama.mcc.ac.uk!bofh.dot!thor.cf.ac.uk!news From: Nick Jacobsen Subject: Re: DNA ligation help Sender: news@cf.ac.uk (USENET News System) Message-ID: Date: Mon, 3 Jun 1996 09:20:40 GMT To: xcl@med.unc.edu X-Nntp-Posting-Host: d053.mg.cf.ac.uk Content-Transfer-Encoding: 7bit Content-Type: text/plain; charset=us-ascii References: <4opneu$nke@newz.oit.unc.edu> Mime-Version: 1.0 X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Organization: Institute of Medical Genetics Cardiff Lines: 14 Xref: biosci bionet.molbio.ageing:2742 bionet.molbio.bio-matrix:730 bionet.molbio.evolution:4574 bionet.molbio.gene-linkage:1078 bionet.molbio.methds-reagnts:45234 Hi, I find that the majority of problems with DNA ligation inefficiency are due to ATP degradation in the reaction buffer. If in doubt order new buffer and freeze in small aliquots. You can also order Pharmacia ligase - you have to order separate 10mM ATP with it to add to the reaction as required. You have to be fairly accurate with the amount of ATP added as too much is as bad as too little. I hope this is of help to you cheers Nick Jacobsen. From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Path: biosci!daresbury!yama.mcc.ac.uk!bofh.dot!thor.cf.ac.uk!news From: Nick Jacobsen Subject: Re: DNA ligation help Sender: news@cf.ac.uk (USENET News System) Message-ID: Date: Mon, 3 Jun 1996 09:20:12 GMT To: xcl@med.unc.edu X-Nntp-Posting-Host: d053.mg.cf.ac.uk Content-Transfer-Encoding: 7bit Content-Type: text/plain; charset=us-ascii References: <4opneu$nke@newz.oit.unc.edu> Mime-Version: 1.0 X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Organization: Institute of Medical Genetics Cardiff Lines: 14 Xref: biosci bionet.molbio.ageing:2741 bionet.molbio.bio-matrix:729 bionet.molbio.evolution:4573 bionet.molbio.gene-linkage:1077 bionet.molbio.methds-reagnts:45233 Hi, I find that the majority of problems with DNA ligation inefficiency are due to ATP degradation in the reaction buffer. If in doubt order new buffer and freeze in small aliquots. You can also order Pharmacia ligase - you have to order separate 10mM ATP with it to add to the reaction as required. You have to be fairly accurate with the amount of ATP added as too much is as bad as too little. I hope this is of help to you cheers Nick Jacobsen. From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Path: biosci!daresbury!yama.mcc.ac.uk!bofh.dot!thor.cf.ac.uk!news From: Nick Jacobsen Subject: Re: DNA ligation help Sender: news@cf.ac.uk (USENET News System) Message-ID: Date: Mon, 3 Jun 1996 09:19:49 GMT To: xcl@med.unc.edu X-Nntp-Posting-Host: d053.mg.cf.ac.uk Content-Transfer-Encoding: 7bit Content-Type: text/plain; charset=us-ascii References: <4opneu$nke@newz.oit.unc.edu> Mime-Version: 1.0 X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Organization: Institute of Medical Genetics Cardiff Lines: 14 Xref: biosci bionet.molbio.ageing:2740 bionet.molbio.bio-matrix:728 bionet.molbio.evolution:4572 bionet.molbio.gene-linkage:1076 bionet.molbio.methds-reagnts:45232 Hi, I find that the majority of problems with DNA ligation inefficiency are due to ATP degradation in the reaction buffer. If in doubt order new buffer and freeze in small aliquots. You can also order Pharmacia ligase - you have to order separate 10mM ATP with it to add to the reaction as required. You have to be fairly accurate with the amount of ATP added as too much is as bad as too little. I hope this is of help to you cheers Nick Jacobsen. From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Newsgroups: bionet.molbio.ageing,bionet.molbio.bio-matrix,bionet.molbio.evolution,bionet.molbio.gene-linkage,bionet.molbio.gene-org,bionet.molbio.methds-reagnts Path: biosci!daresbury!yama.mcc.ac.uk!bofh.dot!thor.cf.ac.uk!news From: Nick Jacobsen Subject: Re: DNA ligation help Sender: news@cf.ac.uk (USENET News System) Message-ID: Date: Mon, 3 Jun 1996 09:20:58 GMT To: xcl@med.unc.edu X-Nntp-Posting-Host: d053.mg.cf.ac.uk Content-Transfer-Encoding: 7bit Content-Type: text/plain; charset=us-ascii References: <4opneu$nke@newz.oit.unc.edu> Mime-Version: 1.0 X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Organization: Institute of Medical Genetics Cardiff Lines: 14 Xref: biosci bionet.molbio.ageing:2743 bionet.molbio.bio-matrix:731 bionet.molbio.evolution:4575 bionet.molbio.gene-linkage:1079 bionet.molbio.methds-reagnts:45235 Hi, I find that the majority of problems with DNA ligation inefficiency are due to ATP degradation in the reaction buffer. If in doubt order new buffer and freeze in small aliquots. You can also order Pharmacia ligase - you have to order separate 10mM ATP with it to add to the reaction as required. You have to be fairly accurate with the amount of ATP added as too much is as bad as too little. I hope this is of help to you cheers Nick Jacobsen. From owner-bio-matrix@net.bio.net Sun Jun 02 23:00:00 1996 Path: biosci!bcm.tmc.edu!pendragon!news.msfc.nasa.gov!newsfeed.internetmci.com!in2.uu.net!news3.agis.net!agis!atmnet.net!usenet From: Larry Richard Newsgroups: bionet.molbio.bio-matrix Subject: JAVA: HEATING UP THE NET - DEMONSTRATIONS AND APPLICATIONS Date: Mon, 03 Jun 1996 14:57:21 -0700 Organization: OnLine Expo 96 Lines: 7 Message-ID: <31B35FC1.7157@aol.com> NNTP-Posting-Host: 204.250.83.43 Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 2.01Gold (Win95; I) These and other exciting new programming tools are allowing your company to produce creative and dynamic environments with which to get your message across to your online customers. This is a must attend session for companies that want to have a first hand look at the industry's hottest new applications. ONLINE EXPO '96 AT SAN FRANCISCO EXPOSITION CENTER JULY 11-13 (http://www.onlineexpo.com) --email: olexpo96@aol.com From owner-bio-matrix@net.bio.net Tue Jun 11 23:00:00 1996 Path: biosci!TIAC.COM!cathyom From: cathyom@TIAC.COM ("Catherine O'Malley") Newsgroups: bionet.molbio.bio-matrix Subject: Shareware or Freeware for Science Date: 12 Jun 1996 09:27:51 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 12 Sender: daemon@net.bio.net Distribution: world Message-ID: <31BEEEB2.761@tiac.com> NNTP-Posting-Host: net.bio.net Do you know of shareware or freeware that could assist biochemists, molecular biologists or pharmacologists? If so, please tell me how this software can be obtained. I am constructing a home page to facilitate the efforts of working scientists who want to locate tools for common tasks. Links to purely educational software would also be appreciated. At this time, I am only interested software that can be run under Windows. Thanks a lot. Your response will help fellow researchers save valuable time. It's amazing what can be accomplished if we help one another! Catherine McKeon-O'Malley From owner-bio-matrix@net.bio.net Fri Jun 14 23:00:00 1996 Path: biosci!internet!biosci!not-for-mail From: biohelp (BIOSCI Administrator) Newsgroups: bionet.molbio.bio-matrix Subject: IMPORTANT - BIOSCI Fundraising Update! Date: 15 Jun 1996 02:00:33 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 154 Sender: daemon@net.bio.net Distribution: world Message-ID: <199606150900.CAA26062@net.bio.net> NNTP-Posting-Host: net.bio.net BIOSCI is about halfway to its funding goal!! I'm interrupting the usual monthly posting of the BIOSCI miniFAQ to bring you up to date on BIOSCI fundraising progress, a topic of concern to your future use of this resource. Thank you in advance for taking the time to read this message carefully. Last year we announced that BIOSCI was going to adopt the U.S. Public Broadcasting System model to fund its operations after our DOE/NSF grant runs out later this year. Unlike PBS, we are not soliciting contributions from users; we are only selling ads on our Web pages solely to cover our operating costs. Our goal is to seek sponsorships until we build up an operating reserve of about $100,000 and then cease further promotions until we need to build the reserve back up. (The accountants among our readership will be familiar with the problem of deferred revenue which we can not safely utilize until ads have been displayed for a period of time.) We are only about halfway to our funding goal and need to raise further funds to avoid having to curtail services at net.bio.net. Fundraising is time-consuming, however, and we need your help as explained further below. Our operating costs consist of our network connection, phone lines, hardware maintenance (we will be getting newer and faster hardware soon!), plus 0.7 FTE of salaries covering UNIX systems admin, technical support, quality assurance, i.e., testing, of our system, and administrative costs (such as the time it takes to actually find/write/call potential sponsors and raise money!). Although the BIOSCI staff does get compensated for a portion of the work that they do, this project has always received a lot of free after-hours and "vacation" time labor, so we hope that no one will begrudge the time that we do charge to the project to serve you. All of the three part-time staff members, Dave Mack, Julie Lawrence, and myself, have full time day jobs and families in addition to working hard to keep this service running for all of you. Julie and Dave Mack are subcontractors for BIOSCI; my time that is charged to the project defrays a portion of my regular salary instead of adding to my income. Besides having to relocate the project, we were very busy this last year building new infrastructure such as our WWW hypermail interface to the system. This was released last December along with scores of WAIS indices for the newsgroups. Virtually everything is complete, although we do continue to find and fix bugs (many through your helpful feedback!). We are still having some problems with our WAIS indexing. The archives continue to grow rapidly. We are running over 100 indexes now versus three previously and any systems crashes cause greater havoc with the indexing than before! We are still working to fix this as fast as our resources permit and appreciate your patience, but we have been able to automate a lot of the infrastructure to reduce labor as compared to past requirements. We have also implemented new software to make moderation of BIOSCI/bionet newsgroups much easier and combat the growing problem of Internet junk mail and USENET "spamming." About 20% of our groups are now moderated, many of them by the BIOSCI staff! This, for example, made a major difference last year in the quality of content in our EMPLOYMENT/bionet.jobs.offered newsgroup which many commercial concerns and recruiting firms are using **without charge** to recruit candidates for positions in the biological sciences. We are also now in a position to have sponsors for individual newsgroups as you will have noticed if you have visited http://www.bio.net/ and clicked on "Access the BIOSCI/bionet newsgroups" recently. So, how can you help?? ---------------------- As noted above it can take a lot of time to contact potential sponsors if I have to do it all myself. Our request is quite simple. You can do two important things which will take very little time for you individually. First, please use our WWW system at http://www.bio.net/ to access the archives. You can now post or reply to messages via your Web browser. Your usage helps attract sponsors. If you contact any of our sponsors, please be sure to thank them for supporting BIOSCI. It is critical for them to get this feedback if they are to continue their sponsorship for the long term. Second, if you work for a company or organization that provides products or services of interest to the biology community, please pass this message on to your marketing or marketing communications department or other appropriate group. Please ask them to help support BIOSCI by sponsoring our Web site and explain the uses and benefits of the system to the biology community. If they are interested, they can then contact us for further information at our tech support address, biosci-help@net.bio.net. Our hope is to quickly raise several large corporate/institutional sponsors on our heavily-used WWW locations (some stats appended below), and then end this sponsorship campaign so that our resources can continue to be used for service provision, not fundraising. Many of our specialty newsgroup WWW archives are still used by small communities of scientists (and they haven't been heavily promoted yet). While these may be valuable niche markets to some advertisers, it will generate more labor and overhead having to find these sponsors, fairly price the locations, and deal with lots of smaller sponsorships than fewer mid-to large sponsors. We are striving to keep our operation as lean and efficient as possible since we are not trying to make careers out of running BIOSCI. We are trying if at all possible to avoid the administrative overhead entailed with processing lots of small payments to reach our fundraising goals. I'd like to thank all of you for your help in advance. In helping us, you are also helping yourselves, not only in keeping this resource available for all of the both large and small research communities that we serve, but also by alleviating the need for us to go back and compete with researchers for tight grant dollars! We promised NSF when we were awarded the BIOSCI grant that we would carry out this mission to make the service self-supporting. With your help, we will succeed in continuing BIOSCI's work into its second decade. Thank you very much! Sincerely, Dave Kristofferson BIOSCI/bionet Manager biosci-help@net.bio.net A list of our prime WWW sponsorship locations follow. Please contact us for further details. ---------------------------------------------------------------------- The overall BIOSCI WWW pages are currently visited by users from close to 5500 unique computer hosts per week. Web servers only log the Internet computer/host name and frequently more than one individual can connect to us from a particular host. Main home page, http://www.bio.net, visited recently by about 2100 unique hosts per week Main Newsgroups archives page, http://www.bio.net/archives.html, visited recently by about 1200 Unique hosts per week BIO-JOURNALS archive page, http://www.bio.net/BIO-JOURNALS.html, visited recently by about 1000 unique hosts per week. EMPLOYMENT archive pages: http://www.bio.net:80/hypermail/EMPLOYMENT/ and monthly header pages, visited recently by about 800 unique hosts per week. Address database search page, http://www.bio.net/addrsearch.html, visited recently by about 450 unique hosts per week. Methods newsgroup archive pages, http://www.bio.net:80/hypermail/METHDS- REAGNTS/ and monthly header pages, visited recently by about 350 unique hosts per week. Ads can also be displayed on various combinations of other BIOSCI/bionet newsgroups. Please contact us at biosci-help@net.bio.net for details. ---------------------------------------------------------------------- From owner-bio-matrix@net.bio.net Thu Jun 27 23:00:00 1996 Path: biosci!ynu.edu.cn!hongluo From: hongluo@ynu.edu.cn (hongluo) Newsgroups: bionet.molbio.bio-matrix Subject: helpal Date: 28 Jun 1996 05:46:35 -0700 Organization: Yunnan University Lines: 1 Sender: daemon@net.bio.net Distribution: world Message-ID: <31D46F69.36D6@ynu.edu.cn> NNTP-Posting-Host: net.bio.net Help. From owner-bio-matrix@net.bio.net Thu Jun 27 23:00:00 1996 Path: biosci!bcm.tmc.edu!cs.utexas.edu!usc!chi-news.cic.net!news.compuserve.com!news.production.compuserve.com!news From: Pierre Dion <102605.2065@CompuServe.COM> Newsgroups: aus.mbio,bionet.molbio.bio-matrix,bionet.molbio.genbank,bionet.molbio.gene-linkage,sci.research.careers Subject: Molecular/Cellular RESEARCH-TORONTO Date: 28 Jun 1996 15:40:42 GMT Organization: Dion Management Consulting Group Lines: 24 Message-ID: <4r0udq$4dt$1@mhade.production.compuserve.com> Xref: biosci bionet.molbio.bio-matrix:740 bionet.molbio.genbank:2319 bionet.molbio.gene-linkage:1100 sci.research.careers:11051 We have recently been mandated by a prominent Canadian biotechnological company to recruit a person to head their molecular and cellular research. Our client's mission is focused upon discovering and developing innovative therapeutics using modern molecular and chemical technologies. Significant disease targets are identified through advanced biological techniques. Strategic alliances are an integral part of corporate strategy for moving in-house technologies to commercialization. The incumbent will report to the senior Vice-President, Research and Clinical Development, and will directly supervise five to six people and indirectly between twenty-five to thirty-five people. We are looking for people with a Ph.D. in Molecular Biology/Biochemistry and at least 8 years research experience in an industrial environment following postdoctoral training in relevant experimental areas in a multidisciplinary environment. This position is located in Toronto with an attractive compensation package. If you are interested in submitting your resume or obtaining more information about this position, please contact Marlene Harris at 800 261-3204. -- Dion From owner-bio-matrix@net.bio.net Fri Jun 28 23:00:00 1996 Path: biosci!rutgers!uwm.edu!vixen.cso.uiuc.edu!news.uoregon.edu!hunter.premier.net!uunet!inXS.uu.net!server-b.cs.interbusiness.it!qualcuno.nettuno.it!sirio.cineca.it!gopher From: Marco Radice Newsgroups: bionet.molbio.bio-matrix Subject: Mesenchymal stem cells Date: 28 Jun 1996 21:54:28 GMT Organization: Institute of Histology - University of Padova Lines: 19 Message-ID: <4r1kak$7l0@sirio.cineca.it> NNTP-Posting-Host: ppp2.dei.unipd.it Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.22 (Windows; I; 16bit) Hi dear netters, I am doing some work on mesenchymal stem cells and I am wondering if anybody out there is trying to isolate those elements from the peripheral circulating blood. Are particular growth factors necessary for a better recovery from blood of those cells? Please answer directly to me if you do not mind. Thanks in advance. Marco Radice Inst. of Histology Univ. of Padova