From owner-autoseq@hgmp.mrc.ac.uk Sun Jul 1 13:19:40 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 4FDFB17A72 for ; Sun, 1 Jul 2001 13:19:39 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 76A6F17A89 for ; Sun, 1 Jul 2001 13:19:37 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 25FB317A72; Sun, 1 Jul 2001 13:19:34 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 29 Jun 2001 19:25:41 +0100 From: SMarsh3807 Subject: Re: ABI 3100: red dye blob at 240 basecalls Message-Id: <20010701121934.25FB317A72@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk One thing to becareful of is missing part of the column when doing the cleanup. I have seen this myself on both our 3100 and 3700. Sometimes loading hte column too fast will cause some sample to run down the side without gong through the column. We stopped using the G50 columns as they didn't work as well (we saw more blobs and weaker signal) so we switched to the Qiagen Dye-Ex removal 96 well plates. they work very well for us. The one thing we have noticed is that you have to wash them after you remove the buffer before loading the samples. WE run 200 ul of water through then load the samples. WE are getting much brighter signals with the BD v3.0 since we started washing the wells. Steve Marsh Director DNA Sequencing Core Facility at Caltech (former ABI service engineer) --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 3 09:43:44 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 6D29E17B2F for ; Tue, 3 Jul 2001 09:43:35 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 54DFE17B1C for ; Tue, 3 Jul 2001 09:43:21 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 1A3E317B49; Tue, 3 Jul 2001 09:38:02 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 3 Jul 2001 03:00:50 +0100 From: KDD 2001 Subject: KDD-2001 Call for Participation Message-Id: <20010703083802.1A3E317B49@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk KDD-2001 Call for Participation The Seventh ACM SIGKDD International Conference on Knowledge Discovery and Data Mining KDD-2001 August 26-29, 2001, San Francisco, CA, USA Register online at http://www.acm.org/sigkdd/kdd2001/ KDD-2001, the premier annual data mining conference organized by the scientists and practitioners who popularized data mining in the mid 1990's, is a must-attend conference, featuring state-of the art keynote talks, free tutorials by experts from academia and industry, rigorously selected technical and industrial papers, the KDD-Cup data mining competition, and more. With over 900 attendees from 34 countries, KDD-2000 held in Boston last year was a great success, bringing together the top minds from academia and the leading industrial practitioners to allow the two groups to share insights and educate attendees who were newer to data mining. Roughly 40% of attendees were from academia, 60% from business. Highlights of the conference this year include: * Keynote presentations by leaders in the field on collaboration, knowledge discovery in biology, and more! * Industrial track invited talks on mining e-commerce data, recommendation systems, predictive modeling, and more * Tutorials -- free with the registration! -- featuring e-business, frequent-pattern mining, outlier analysis, value-based mining, mobile data mining, and advances in decision trees * KDD-Cup 2001: results from this year's genomics and drug design competition * Workshops on visualization, multimedia, scientific data, weblog analysis, bioinformatics, and temporal data * Panels on sampling, data mining startup companies, and new research directions * Exhibits from over 26 leading software & hardware vendors and academic projects * Technical paper presentations from 20 authors, plus 32 poster presentations giving attendees a chance to talk directly with the authors * Industry track papers presented by 12 teams from leading companies such as Microsoft, IBM, Verizon, and others -- learn how the most successful companies put data mining into practice! See the conference schedule and full details at http://www.acm.org/sigkdd/kdd2001 and register online by following the links from the conference homepage. Register by July 27 to save over $100, and be sure to book hotel rooms early for discounted rates! With the first-rate program summarized above, there is something for everyone at KDD-2001 and you won't want to miss this once-a-year opportunity to meet and share knowledge with your data-mining colleagues from around the world. Haipeng Guo, www.kddresearch.org Computing and Information Sciences Department, Kansas State University. --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 3 09:46:07 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 78FAD17B38 for ; Tue, 3 Jul 2001 09:45:59 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 87D3017B55 for ; Tue, 3 Jul 2001 09:45:45 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id E909917AE9; Tue, 3 Jul 2001 09:40:13 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 3 Jul 2001 09:02:17 +0100 From: Roland.Varecka@vie.boehringer-ingelheim.com Subject: Re: ABI 3100: red dye blob at 240 basecalls Message-Id: <20010703084013.E909917AE9@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear Angela, dear Steve, thanks for your hints on loading the multiscreen plates. I ran some test yesterday and the trend is clear: either reducing the loading volume from 30uL to 15u= L (plus making sure the samples are applied exactly at center column) or increasing the amount of Sephadex G-50 from 45uL to 90uL produced very nice and clean sequences.= =20 The latter method gave more consistent sample quality and samples are more easily loaded when using an 8-channel pipettor, however may be a bit messy for routine application, as the swollen gel slightly protrudes over the rim of the wells (reminded m= e a bit of freshly tapped draft beers). I have also ordered the Quiagen DyeEx 96 kit. Thanks again, Sincerly Roland --------------------------------------------------------------- Roland M Varecka Boehringer Ingelheim Austria GmbH Dr. Boehringergasse 5-11 A-1121 Wien, =D6sterreich Tel:=09xx43-1-80105-2392 Fax:=09xx43-1-80105-2360 Email:=09roland.varecka@vie.boehringer-ingelheim.com --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 3 15:16:07 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 13BC517A78 for ; Tue, 3 Jul 2001 15:16:06 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 43B8917AA8 for ; Tue, 3 Jul 2001 15:16:02 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id EDB7A17A78; Tue, 3 Jul 2001 15:15:50 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 3 Jul 2001 14:11:10 +0100 From: Phillip San Miguel Subject: Re: ABI 3100: red dye blob at 240 basecalls Message-Id: <20010703141550.EDB7A17A78@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Roland.Varecka@vie.boehringer-ingelheim.com wrote: > Dear Angela, dear Steve, > > thanks for your hints on loading the multiscreen plates. I ran some test > yesterday > and the trend is clear: either reducing the loading volume from 30uL to 15uL > (plus making sure the samples are applied exactly at center column) or increasing > the amount of Sephadex G-50 from 45uL to 90uL produced very nice and clean sequences. > The latter method gave more consistent sample quality and samples are more > easily loaded when using an 8-channel pipettor, however may be a bit messy for routine application, > as the swollen gel slightly protrudes over the rim of the wells (reminded me > a bit of freshly tapped draft beers). I have also ordered the Quiagen DyeEx > 96 kit. > [...] Actually, of the 3 types of commercial clean-up plates we have used: Qiagen, BioRad and Edge; Qiagen uses the least amount of column material. In our hands BioRad and Edge gave superior results. Edge, especially seemed to be able to completely remove large amounts of unincorporated dye-terminators. Phillip SanMiguel Purdue Genomics Core Facility --- From owner-autoseq@hgmp.mrc.ac.uk Wed Jul 4 07:30:13 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 24B5717B2A for ; Wed, 4 Jul 2001 07:30:10 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id B716D17A5B for ; Wed, 4 Jul 2001 07:30:07 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 030B117A68; Wed, 4 Jul 2001 07:30:01 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 3 Jul 2001 19:36:34 +0100 From: Jarmo Korkko Subject: DyeTerminator removal for Capillary Sequencer (ABI3100) - Edge Message-Id: <20010704063001.030B117A68@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hello, We have been using pre-packed Edge Biosystems DyeTerminator Removal 96-well plates for 7-8 months with really good results (and probably go back to them again). Unfortunately the cost is quite high and therefore we tried "do-it-yourself" Millipore MultiScreen-HV Filter plates with Sephadex G-50 Superfine (~$15 vs $50). We are using the same well-working platform (ABI 3100) to analyze the samples in both cases. We have not been able to get the Millipore system to work well consistently with our ABI 3100 Sequencer. Some samples are fine, some leak dyeterminators (and/or other reagents) really badly. We have tried all the suggested options for rinsing and pre-wetting etc., have tried different centrifugation times. Constantly our signal strenghts are really high (which is probably good or is it?). We have diluted the samples in regards to reaction mix and DNA template which seems to help a little bit but we still end up running some of the samples again after re-diluting them. Does anyone have other ideas? Our situation is complicated by the fact that our samples come from a number of people (core lab) and the sample difficulties seem to be lab specific. That did not use to be the case with Edge plates (if sample was to work the results were nice). Now we are experiencing premature loss of resolution, broad labeled nucleotide(?) peaks in the beginning (first 20-30-bp), signal strentghts are out of control (up to 5000!!! with 1:10 diluted reactions [~300-bp PCR products) and unpredictable. I don't know if the signal is coming from dyeterminators that didn't get removed or from other reagents that weren't removed in the process. Or do the Edge columns really bind that much perfectly good product in their columns? Is the loss of resolution from too much DNA trying to go through the capillary or are salt etc remnants from the sample causing the trouble. Edge plates have higher columns than Millipore, that might play a role.. We have decent results with our DNA robot when samples are quite diluted and homogeneous, still consistenly some peaks in the first 20-30-bp. Any comments are greatly appreciated. Thanks, Jarmo Jarmo Korkko, M.D., Ph.D. Research Assistant Professor Tulane University Health Sciences Center Center for Gene Therapy 1324 Tulane Avenue, SL99 New Orleans, LA 70112 Phone intl+1-504-988-7708 Cell Phone 504-388-3436 Fax 504-988-7704 --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 5 17:45:51 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 2766F17AE7 for ; Thu, 5 Jul 2001 17:45:48 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 3A66717AEE for ; Thu, 5 Jul 2001 17:45:42 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 955B617AE7; Thu, 5 Jul 2001 17:45:32 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 5 Jul 2001 14:48:34 +0100 From: "PBO (Peter Bjarke Olsen)" Subject: Re: DyeTerminator removal for Capillary Sequencer (ABI3100) - Edg e Message-Id: <20010705164532.955B617AE7@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi Jarmo, We use the Milliporesystem with good results. We do the following. 1:Add 300 ul water 2: Let the sephadex swell for 60 minutes. 3: Centrifuge 3 min at 1900Xg 4: Add 200 ul water 5: Centrifuge 3 min at 1900xg 6: Add sample and centrifuge 3 min at 1900xg . 7: The samples are loaded directly. We dilute the bigdye terminitors (v2) 4 times and we do 50 pcr cycles. In general we see only small amounts of dye blobs. As other have metioned it is important to load the sample in the middle of the wells. >From may to june 2001 we produced 14232 sequences (54 different users or projects). Average phred20: 490 Highest phred20 score: 788 Lowest phred20 score: 0 Average signal: 267 Highest signal: 3023 lowest signal 2.75 We use a 3700 so I don't know if the signals can be compared. Has anyone tried to use this system (or a similar gelfiltration system) with version 3 of the Bigdyes? Applied Biosystems have told me that it doesn't work. Best regards, Peter Bjarke Olsen --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 6 17:27:39 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id DCE7F17A78 for ; Fri, 6 Jul 2001 17:27:35 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id BDA8117A84 for ; Fri, 6 Jul 2001 17:27:32 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id BADAB17A78; Fri, 6 Jul 2001 17:27:25 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 6 Jul 2001 16:38:33 +0100 From: Darek Subject: Ethanol precipitation with 3100? Message-Id: <20010706162725.BADAB17A78@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi Guys, We just got new 3100, so we will be switching from 373 to 3100. We've used modified ethanol precipitation protocol to clean samples on 373. We'tried a couple of commercial clean-up plates but our method worked best. The question is: does anyone have an experience with using ethanol precipitation on 3100? regards DArek --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 9 09:32:27 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 864EB17ABB for ; Mon, 9 Jul 2001 09:32:25 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id C810D17AD9 for ; Mon, 9 Jul 2001 09:32:20 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id E46DF17ABB; Mon, 9 Jul 2001 09:32:16 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 9 Jul 2001 08:13:12 +0100 From: Bernd Weisshaar Subject: DNAseq: DyeTerminator removal for Capillary Sequencer Message-Id: <20010709083216.E46DF17ABB@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear DNA-Sequers, dear Peter, below, there is our modification of the millipore purification procedure which works well in our hand (it is really close to the original millipore protocol). We are just moving to BDv3.0 for the 3100, and so far the protocol seems to work as good as with v2.0. We may be able to use a little less kit per reaction, but we have not yet tested that in detail. For a 10 ul cycle-sequencing reaction with 2 or 3 ul BD2 (depending on the size of the DNA template): 1: Add 300 ul water 2: Let the sephadex swell for round about 3h. (we use G50 from Pharmacia which is loaded dry into the millipore setup so that the "colums" are filled completely) 3: Centrifuge 5 min at 900 g 4: Add 150 ul water (using a Hydra) 5: Centrifuge 5 min at 900 g 6: Add 9 ul water to the samples, load them **to the centers** of the columns and centrifuge 5 min at 900 g directly into the 96-er MircoAmp Optical Plates which contain 15 ul water/well (we use only HPLC grade water from Merck). We only very occasionally see dye blobs, and I think that in this case the problem almost always is that the loading of the diluted sample was not done carefully to the center of the sephadex, or column loading with sephadex was not OK. The dilution helped in our hands to get constant volumes out of the plates. For the 3700, we "tape" the MicroAmp plates (but no v3.0 experience yet on the 3700 - the service just upgraded the instrument). Best regards, Bernd ======================================================== Hi Jarmo, We use the Milliporesystem with good results. We do the following. 1: Add 300 ul water 2: Let the sephadex swell for 60 minutes. 3: Centrifuge 3 min at 1900Xg 4: Add 200 ul water 5: Centrifuge 3 min at 1900xg 6: Add sample and centrifuge 3 min at 1900xg . 7: The samples are loaded directly. We dilute the bigdye terminitors (v2) 4 times and we do 50 pcr cycles. In general we see only small amounts of dye blobs. As other have metioned it is important to load the sample in the middle of the wells. >From may to june 2001 we produced 14232 sequences (54 different users or projects). Average phred20: 490 Highest phred20 score: 788 Lowest phred20 score: 0 Average signal: 267 Highest signal: 3023 lowest signal 2.75 We use a 3700 so I don't know if the signals can be compared. Has anyone tried to use this system (or a similar gelfiltration system) with version 3 of the Bigdyes? Applied Biosystems have told me that it doesn't work. Best regards, Peter Bjarke Olsen --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 9 10:35:34 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 0563D17A65 for ; Mon, 9 Jul 2001 10:35:27 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 66DFC17A60 for ; Mon, 9 Jul 2001 10:35:10 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 72B2B17A5B; Mon, 9 Jul 2001 10:34:27 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 9 Jul 2001 09:47:10 +0100 From: "Tarragona, Tony" Subject: RE: Ethanol precipitation with 3100? Message-Id: <20010709093427.72B2B17A5B@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk It is not a problem for us, we use the ethanol precipitation adapted for microtiter plates and we get very good results, hardly any dye blobs. Best Toni > ---------- > From: Darek > Sent: Friday, July 6, 2001 16:38 PM > To: autoseq@net.bio.net > Subject: Ethanol precipitation with 3100? > > Hi Guys, > We just got new 3100, so we will be switching from 373 to 3100. > We've used modified ethanol precipitation protocol to clean samples > on 373. We'tried a couple of commercial clean-up plates but our > method worked best. The question is: does anyone have an > experience with using ethanol precipitation on 3100? > regards > DArek > > --- > > --- From owner-autoseq@hgmp.mrc.ac.uk Wed Jul 11 20:17:15 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id D1BA217B54 for ; Wed, 11 Jul 2001 20:17:11 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 85E0517B18 for ; Wed, 11 Jul 2001 20:17:09 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 5EB4317AE5; Wed, 11 Jul 2001 20:17:04 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 11 Jul 2001 19:26:25 +0100 From: John Weger Subject: Job Opening: Research Technician Position at the Ludwig Institute Message-Id: <20010711191704.5EB4317AE5@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Research Technician Position at the Ludwig Institute for Cancer Research, San Diego Branch (UCSD Campus, La Jolla) Brief description- Working in a DNA sequencing and genotyping core facility developing and implementing procedures for high throughput and routine sequencing and genotyping as well as developing and implementing high throughput genetics methods and interfacing them with the facility. Duties- The major duties of this position will be to implement sequencing and genotyping protocols as well as high throughput molecular biology and genetics methods on robotic workstations. These include dye primer and dye terminator sequencing protocols, PCR product generation for sequencing based mutation detection and microarray projects and fluorescent primer based genotyping. The types of projects will include DNA sequencing, large-scale sequencing, genotyping for positional cloning and clinical/population genetics projects and yeast genetics projects. Responsibilities will also include management of projects including high throughput projects, development and optimization of primers and organization, and analysis and reporting of data. Education and experience required- A minimum of a BS/BA degree with a science major, preferably in the Biological Sciences such as genetics, molecular biology or biochemistry. Up to two to three years of experience operating ABI sequencers and knowledge of dye primer and dye terminator chemistry and use of Robotic workstations such as Tecan robots would receive preference. Interest in robotic workstations and other types of complex laboratory instrumentation is a positive attribute. Working knowledge of Macintosh computers including use of database, spreadsheet, statistics, DNA/protein sequence analysis, and genome database search and analysis programs is required. Experience with other types of computer systems will be helpful. Please contact: John Weger Ludwig Institute for Cancer Research UC San Diego School of Medicine, CMME 3031 9500 Gilman Drive La Jolla, CA 92093-0660 jweger@ucsd.edu --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 12 16:13:50 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id D90F417B7B for ; Thu, 12 Jul 2001 16:13:44 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 0119017B2D for ; Thu, 12 Jul 2001 16:13:35 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 9E7CB17B74; Thu, 12 Jul 2001 16:12:53 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 12 Jul 2001 15:06:25 +0100 From: Tony Subject: Phred Quality analysis Message-Id: <20010712151253.9E7CB17B74@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Has anyone out there has any sucess storied with long term monetering of sequence quality using the phred<20 scores for the 3700? I am currently trying to get a quality system based on phred scores internally validated and I would like to hear from people who have done the same thing and how they got on. Amongst my many problems are assigning cut-offs. Thankyou in advance --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 13 09:04:49 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 505D217A5B for ; Fri, 13 Jul 2001 09:04:47 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id A583B17A92 for ; Fri, 13 Jul 2001 09:04:43 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 4A8D317A5B; Fri, 13 Jul 2001 09:04:39 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 12 Jul 2001 23:18:55 +0100 From: Secuenciador Subject: Secuencing facilities Message-Id: <20010713080439.4A8D317A5B@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi, I am looking for information of labs with an automated secuencing comercial service. Any information on characteristics of the templates, primers, equipments used and costs will be very well received. Thenks for your time, Maria --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 13 09:06:48 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 8DACD17AB9 for ; Fri, 13 Jul 2001 09:06:43 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 6E83A17AC0 for ; Fri, 13 Jul 2001 09:06:39 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 13F4317AB9; Fri, 13 Jul 2001 09:06:10 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 13 Jul 2001 08:27:16 +0100 From: Sean Moore Subject: CEQ2000XL seq reaction buffer Message-Id: <20010713080610.13F4317AB9@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear Sequencers, I am using a Beckman CEQ2000XL capillary sequencer with the CEQ DTCS kit. I have had success diluting the sequencing reactions by as much as 1/6 and was wondering if anyone else had a similar experience. If so, is there an alternative to the quite expensive proprietary buffer for use in the dilutions. Any advice would be helpful. Regards, Sean Moore Food Science Australia Sneydes Road Werribee, Melbourne Australia, 3030 sean.moore@foodscience.afisc.csiro.au Phone:+61 3 9731 3313 (Office) +61 3 9731 3296 (Lab) FAX: +61 3 9731 3254 Food Science Australia; a joint venture of CSIRO and Afisc Results for today; ideas for tomorrow. --- From owner-autoseq@hgmp.mrc.ac.uk Sat Jul 14 17:38:43 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 902F717B0A for ; Sat, 14 Jul 2001 17:38:41 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id ACAD117B14 for ; Sat, 14 Jul 2001 17:38:39 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 91A4D17B0A; Sat, 14 Jul 2001 17:38:36 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 13 Jul 2001 22:54:10 +0100 From: Malcolm Cook Subject: Applescripting ABI Prism 377XL Collection software Message-Id: <20010714163836.91A4D17B0A@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk H'lo! Regarding 'ABI Prism 377XL Collection' software version 2.6, I read with interest in http://www.appliedbiosystems.com/support/download/ga/377/ub/pdf/4313687.pdf the following: "Applescript=AE support This application now supports an Apple event that l= ets you import a text file. The information from the text file is used to fill out a sample sheet and, optionally, a run sheet based on the imported sampl= e sheet." However, I've not been able to make it work (yet) and my requests for support (GALAB@appliedbiosystems.com =3D The Genetic Analysis Technical Support Group, Applied Biosystems) come up dry Any one out there got it working? Willing to share their snippet of applescript with me? Thanks, Malcolm Cook (858) 623-7654 BioInformatics DNA Science Laboratories --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 17 14:02:51 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 582CD17AF6 for ; Tue, 17 Jul 2001 14:02:48 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id C136617AD7 for ; Tue, 17 Jul 2001 14:02:45 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id E39D617A81; Tue, 17 Jul 2001 14:02:41 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 17 Jul 2001 11:39:34 +0100 From: Mark T Jung Subject: Re: Phred Quality analysis Message-Id: <20010717130241.E39D617A81@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Tony, We have been using phred for about a year now to QC our sequencing results and have found this to to be very useful. My impression is that most others would agree with this. We did a direct comparison a few months ago with Trace Tuner (Paracel) and found we liked the phred quality assignments better. Paracel claims a 10% greater read length on 3700 data; we could not confirm this on our 3700 data. We use phred's trimming feature to trim our data. E-Mail me directly if you want specifics regarding how we do the analysis. Mark Jung Tony on 07/12/2001 10:06:25 AM To: autoseq@net.bio.net cc: (bcc: Mark T Jung/AE/DuPont) Subject: Phred Quality analysis Has anyone out there has any sucess storied with long term monetering of sequence quality using the phred<20 scores for the 3700? I am currently trying to get a quality system based on phred scores internally validated and I would like to hear from people who have done the same thing and how they got on. Amongst my many problems are assigning cut-offs. Thankyou in advance --- --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 17 16:26:39 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id A0AAB17B8F for ; Tue, 17 Jul 2001 16:26:32 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 64B2F17B75 for ; Tue, 17 Jul 2001 16:26:23 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id A236117B97; Tue, 17 Jul 2001 16:17:30 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 17 Jul 2001 15:17:41 +0100 From: "Osborne, Brian" Subject: RE: Phred Quality analysis Message-Id: <20010717151730.A236117B97@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Tony, I can't speak to the 3700, the last work I was involved in with DNA sequencing used 377's and 373's, but we would typically define "high-quality" as a sequence having an average Phred quality of 11 over a window of 15 nucleotides. So the perl script would start at the ends of each read trimming sequence that didn't meet this criterion until it reached high quality sequence. These values were determined empirically, by examining many reads and noting the Phred scores of what scientists considered legible and illegible sequence. I notice that Mark talks about phred's trimming feature, we didn't use this feature. Brian O. -----Original Message----- From: Mark T Jung [mailto:MARK.T.JUNG@USA.dupont.com] Sent: Tuesday, July 17, 2001 6:40 AM To: autoseq@net.bio.net Subject: Re: Phred Quality analysis Tony, We have been using phred for about a year now to QC our sequencing results and have found this to to be very useful. My impression is that most others would agree with this. We did a direct comparison a few months ago with Trace Tuner (Paracel) and found we liked the phred quality assignments better. Paracel claims a 10% greater read length on 3700 data; we could not confirm this on our 3700 data. We use phred's trimming feature to trim our data. E-Mail me directly if you want specifics regarding how we do the analysis. Mark Jung Tony on 07/12/2001 10:06:25 AM To: autoseq@net.bio.net cc: (bcc: Mark T Jung/AE/DuPont) Subject: Phred Quality analysis Has anyone out there has any sucess storied with long term monetering of sequence quality using the phred<20 scores for the 3700? I am currently trying to get a quality system based on phred scores internally validated and I would like to hear from people who have done the same thing and how they got on. Amongst my many problems are assigning cut-offs. Thankyou in advance --- --- --- From owner-autoseq@hgmp.mrc.ac.uk Wed Jul 18 18:46:14 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id ED64C17B74 for ; Wed, 18 Jul 2001 18:46:12 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 1D0CD17B6C for ; Wed, 18 Jul 2001 18:46:11 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id E969717B74; Wed, 18 Jul 2001 18:46:07 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 18 Jul 2001 17:14:04 +0100 From: Christine Ponte Subject: 3100 and Office 2000 Message-Id: <20010718174607.E969717B74@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Has anyone downloaded Office 2000 on the 3100's NT machine. My NT machine is not on a network and I would like to utilize the printer for other applications specifically graphics on Powerepoint presentations. I was wondering if running the office program interfered with communication with the intrument. Thanks Christine Ponte, M.S. Dept. of Medicine/Division of Hematology/Cardeza 1015 Walnut St., Room 702 Phila. PA, 10107 215-955-1529 office, 215-955-2256 lab --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 09:42:37 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id C770417AAC for ; Thu, 19 Jul 2001 09:42:35 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id EED1117AF3 for ; Thu, 19 Jul 2001 09:42:33 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id BDB5E17AAC; Thu, 19 Jul 2001 09:42:30 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 18 Jul 2001 22:56:56 +0100 From: George S.Grills Subject: positions open at the Harvard-Partners Center for Genetics and Message-Id: <20010719084230.BDB5E17AAC@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk The Harvard-Partners Center for Genetics and Genomics is building a state-of-the-art facility in Cambridge that will be dedicated to high throughput genotyping, DNA sequencing and DNA microarray technology. The Center is currently seeking highly motivated, experienced individuals interested in working in an academically-oriented team environment to fill positions in the following areas: Facility Managers: Ph.D. level candidates having prior experience with high throughput technologies are sought. Industrial or biotechnology background preferred. Responsibilities include overseeing sequencing, genotyping and microarray facilities. Knowledge and experience in personnel management, budgeting and computation are required. Opportunities for research and development of new technologies will be available. Laboratory Technicians: Several full-time B.S. and M.S. level positions requiring proven skills in molecular biology techniques including PCR, DNA purification, nucleic acid hybridization, cloning, and automated DNA sequencing. Excellent computer skills are essential. Bioinformaticists: B.S. through Ph.D. level candidates with demonstrated experience in computational biology. Knowledge of molecular biology is a plus. Systems Analyst: Responsible for maintaining high end computers and data processing of several large-scale genetics programs. Executive Assistant to the Scientific Director: Responsible for assisting the Director in managing all aspects of the Center. Please send or e-mail a statement of interest, resume and list of three references to: Elaine O'Rourke Division of Genetics, Department of Medicine Brigham and Women's Hospital 20 Shattuck Street Boston, MA 02115 eorourke@rascal.med.harvard.edu Brigham and Women's Hospital is an equal opportunity/affirmative action employer. --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 09:45:36 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id A36F717BA7 for ; Thu, 19 Jul 2001 09:45:27 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 2E93117B96 for ; Thu, 19 Jul 2001 09:45:19 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 9D7E717BA9; Thu, 19 Jul 2001 09:43:34 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 19 Jul 2001 07:12:05 +0100 From: SMarsh3807 Subject: Re: 3100 and Office 2000 Message-Id: <20010719084334.9D7E717BA9@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk I installed office on my 3100 and 3700 and have had no problems related to it. Steve Marsh Director DNA Sequencing Core @ Caltech --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 12:14:56 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 71AF617B1E for ; Thu, 19 Jul 2001 12:14:53 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 7586A17AC9 for ; Thu, 19 Jul 2001 12:14:49 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 37B8917A4E; Thu, 19 Jul 2001 12:14:43 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 19 Jul 2001 10:56:59 +0100 From: Joanne Gilfillan Subject: PE biosystems version 3 sequencing kit Message-Id: <20010719111443.37B8917A4E@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk PLEASE HELP!!! We recently tried out the Big Dye Terminator Kit Version 3 on our ABI 377 sequencer. We experienced an offset/shift of the GTP in relation to the other bases, creating problems with the base calling. Version 1 of this kit gave us no problems at all. Has anyone else also found this, and if so, are there any ideas on how to remedy this problem? Does anyone know whether version3 has been tested extensively on gel-based systems? --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 14:28:26 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id EF2E017AB5 for ; Thu, 19 Jul 2001 14:28:22 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9953917B9D for ; Thu, 19 Jul 2001 14:28:19 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 0D4C617B9E; Thu, 19 Jul 2001 14:28:15 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 19 Jul 2001 14:16:15 +0100 From: Glenis Wiebe Subject: Re: PE biosystems version 3 sequencing kit Message-Id: <20010719132815.0D4C617B9E@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi Joanne, I was very glad to see your message. I am having exactly the same problem - "an offset/shift" of bases causing trouble with the base calling. What I see primarily are N's being called where there are no bases; usually between about 200 and 400 bases. It is not a drop in resolution - the peaks are still well defined. I think it is a problem with the mobility file, because if I analyze the sample with the BDv2 mobility file (DT{BD Set Any-Primer}), that region is fine. (Unfortunately, the peak shifts still occur near the beginning of the sequence.) To make things even more complicated, sometimes the problem is not as severe as others. For the record, I always set up duplicates with the BDv2 kit, and those results are "perfect". Sorry, I know this is not a solution, only a conformation that you are not alone. I hope someone has some ideas how to fix this. Glenis Joanne Gilfillan wrote: > PLEASE HELP!!! > > We recently tried out the Big Dye Terminator Kit Version 3 on our ABI > 377 sequencer. We experienced an offset/shift of the GTP in relation > to the other bases, creating problems with the base calling. Version > 1 of this kit gave us no problems at all. Has anyone else also found > this, and if so, are there any ideas on how to remedy this problem? > Does anyone know whether version3 has been tested extensively on > gel-based systems? > > --- -- Glenis Wiebe, M.Sc. DNA Sequencing Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstrasse 108 D-01307 Dresden Germany Tel: +49-351-210-2628 Fax: +49-351-210-2000 e-mail: wiebe@mpi-cbg.de webpage: www.mpi-cbg.de --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 18:03:47 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id B362F17B9B for ; Thu, 19 Jul 2001 18:03:41 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 60C5617B28 for ; Thu, 19 Jul 2001 18:03:32 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 69A8317B9B; Thu, 19 Jul 2001 18:03:05 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 19 Jul 2001 16:02:44 +0100 From: Dr.Duncan Clark Subject: Re: PE biosystems version 3 sequencing kit Message-Id: <20010719170305.69A8317B9B@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk In article <9j6ff2$q6t$1@mercury.hgmp.mrc.ac.uk>, the eminent Joanne Gilfillan at BIOSCI/MRC Human Genome Mapping Project Resource Centre wrote >We recently tried out the Big Dye Terminator Kit Version 3 on our ABI >377 sequencer. We experienced an offset/shift of the GTP in relation >to the other bases, creating problems with the base calling. Version >1 of this kit gave us no problems at all. Has anyone else also found >this, and if so, are there any ideas on how to remedy this problem? >Does anyone know whether version3 has been tested extensively on >gel-based systems? Just a thought. Do you not need to generate a new matrix for the kit, which will then overcome these problems? Duncan -- It might look like I'm doing nothing, but at the cellular level I'm really quite busy. Duncan Clark GeneSys Ltd. Tel. +44 (0) 1252376288 FAX +44 (0) 8701640382 --- From owner-autoseq@hgmp.mrc.ac.uk Thu Jul 19 22:17:03 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9E8E417A69 for ; Thu, 19 Jul 2001 22:17:01 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9CCDD17B03 for ; Thu, 19 Jul 2001 22:16:59 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 43ED317A69; Thu, 19 Jul 2001 22:16:56 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 19 Jul 2001 21:33:24 +0100 From: SMarsh3807 Subject: Re: PE biosystems version 3 sequencing kit Message-Id: <20010719211656.43ED317A69@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk WE have seen a similar problem, but on our 3700. I was happening only if we left the plate for a couple of hours after we cleaned them up. The plates that sat in water we got shifting "c"s to the point where it was almost unreadable. Interestingly if we dried the samples down and resuspended them in Formamide and ran them on the 3100 they did not do this. Steve --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 20 11:12:44 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 3C1B217B63 for ; Fri, 20 Jul 2001 11:12:40 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id F2D8517AEB for ; Fri, 20 Jul 2001 11:12:37 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 0653F17BAF; Fri, 20 Jul 2001 11:12:33 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 20 Jul 2001 07:33:15 +0100 From: Glenis Wiebe Subject: Re: PE biosystems version 3 sequencing kit Message-Id: <20010720101233.0653F17BAF@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi Duncan, Yes, that was a thought I had too - maybe something had gone horribly wrong while generating the new matrix. However, after making three separate ones (two from matrix standards, one from a sample file), I've decided the problem must be something else. Glenis "Dr.Duncan Clark" wrote: > In article <9j6ff2$q6t$1@mercury.hgmp.mrc.ac.uk>, the eminent Joanne > Gilfillan at BIOSCI/MRC Human Genome Mapping Project Resource Centre > wrote > >We recently tried out the Big Dye Terminator Kit Version 3 on our ABI > >377 sequencer. We experienced an offset/shift of the GTP in relation > >to the other bases, creating problems with the base calling. Version > >1 of this kit gave us no problems at all. Has anyone else also found > >this, and if so, are there any ideas on how to remedy this problem? > >Does anyone know whether version3 has been tested extensively on > >gel-based systems? > > Just a thought. Do you not need to generate a new matrix for the kit, > which will then overcome these problems? > > Duncan > -- > It might look like I'm doing nothing, but at the cellular level I'm really > quite busy. > > Duncan Clark > GeneSys Ltd. > Tel. +44 (0) 1252376288 > FAX +44 (0) 8701640382 > > --- -- Glenis Wiebe, M.Sc. DNA Sequencing Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstrasse 108 D-01307 Dresden Germany Tel: +49-351-210-2628 Fax: +49-351-210-2000 e-mail: wiebe@mpi-cbg.de webpage: www.mpi-cbg.de --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 20 16:02:28 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 893F317AE6 for ; Fri, 20 Jul 2001 16:02:26 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 612F917AE1 for ; Fri, 20 Jul 2001 16:02:24 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id C5A6C17AF7; Fri, 20 Jul 2001 16:02:14 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 20 Jul 2001 13:37:00 +0100 From: Fabien Jovelin Subject: genomic sequencing Message-Id: <20010720150214.C5A6C17AF7@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk I need help for genomic direct sequencing. I have tried the bruce Roe protocol with thermofidelase but never succeed in sequencing my genomic dna= , even with sheared (mecanical) dna. Any hint welcome. Thanks -- Fabien Jovelin ESGS-Cyberg=E8ne --- From owner-autoseq@hgmp.mrc.ac.uk Fri Jul 20 21:40:18 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 2D67B17B53 for ; Fri, 20 Jul 2001 21:40:17 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 51D4517B43 for ; Fri, 20 Jul 2001 21:40:15 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 77EDE17B54; Fri, 20 Jul 2001 21:40:10 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 20 Jul 2001 17:38:47 +0100 From: Glenis Wiebe Subject: Re: BigDye version 3 sequencing kit Message-Id: <20010720204010.77EDE17B54@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk To everyone who has been following the discussion surrounding BigDye version 3 problems, I just received the following from AppliedBiosystems. Since this discussion has been split between two newsgroups, let me give a quick summary of the problem first. I am running BigDye version 3 on a 377 using 48cm 5% LongRanger gels with TTE (Tris/TAPS/EDTA). What I have been experiencing is an offset/shift of G's in relation to the other bases (particularly A's) creating problems with the base calling. So anyway, here is the news: > We hope that by next week there is a mobility file posted on our website > that can handle these long read gels. Please try with this one when it is > released. You find it on our website: > www.appliedbiosystems.com > services & support > softwaredownloads > > ABI Prism 377 > module files > I presume it will have "LR" in its naming. > -- Glenis Wiebe, M.Sc. DNA Sequencing Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstrasse 108 D-01307 Dresden Germany Tel: +49-351-210-2628 Fax: +49-351-210-2000 e-mail: wiebe@mpi-cbg.de webpage: www.mpi-cbg.de From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 09:47:12 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 93F3017B8B for ; Mon, 23 Jul 2001 09:47:06 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 8922E17BA1 for ; Mon, 23 Jul 2001 09:46:56 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 880F817B8B; Mon, 23 Jul 2001 09:46:07 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 21 Jul 2001 20:43:18 +0100 From: Tor Slettnes Subject: Re: 3100 and Office 2000 Message-Id: <20010723084607.880F817B8B@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk >>>>> "Christine" =3D=3D Christine Ponte wri= tes: Christine> Has anyone downloaded Office 2000 on the 3100's NT Christine> machine. My NT machine is not on a network and I would Christine> like to utilize the printer for other applications Christine> specifically graphics on Powerepoint presentations. I Christine> was wondering if running the office program interfered Christine> with communication with the intrument. Thanks No, running Office should be fine. =20 It might be best to install it from a CD, if you have one. If you network your computer (via the 2nd ethernet card), even for the short duration of getting an Office suite onto it, you should change the default passwords (for 3100user, DNA, and Administrator) - otherwise anyone will be able to access your 3100 workstation via FTP. -tor --=20 F=E5r i ulvekl=E6r --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 09:47:59 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 4420617BBF for ; Mon, 23 Jul 2001 09:47:45 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 2DC3817BE9 for ; Mon, 23 Jul 2001 09:47:23 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 74E9E17BE8; Mon, 23 Jul 2001 09:46:51 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 22 Jul 2001 03:35:25 +0100 From: TONTO Subject: Sequencing directly from BACs Message-Id: <20010723084651.74E9E17BE8@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Anyone have any tricks for sequencing directly from BAC clones? Our core lab has never done it and they would like some input before they do some runs for us. I believe they are using the big dye terminator kit and a capillary sequencing machine. I can get more specific info if anyone has had luck with this. Thanks, Tonto tontogoldstein@hotmail.com --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 09:52:28 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id DD00217B3A for ; Mon, 23 Jul 2001 09:52:26 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 0A81B17B8B for ; Mon, 23 Jul 2001 09:52:24 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 518E117B3A; Mon, 23 Jul 2001 09:52:21 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 22 Jul 2001 15:01:08 +0100 From: Ant Smith Subject: DNA sequencing review Message-Id: <20010723085221.518E117B3A@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Hi I am in the process of writing a term paper for my MBA on advances in DNA sequencing technologies. Can anyone point me in the direction of a good review or book that gives an overview of advances in DNA sequencing technologies from the very first methods, through radioactive manual sequencing to automated sequencing. My particular interest is to be able to plot increases in run length and accuracy over the years and correlate this to changes in the technology used. In addition, I would like to try to find a plot of $ per base cost over time. Any suggestions? -- Ant Smith (PhD) Complete Outsourcing ant@completeoutsourcing.com www.completeoutsourcing.com Tel: 083 414 0550 Fax: 083 8 414 0550 From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 15:55:12 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9D71A17B8F for ; Mon, 23 Jul 2001 15:55:08 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 8C7C817ACA for ; Mon, 23 Jul 2001 15:55:01 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id CDFED17B8F; Mon, 23 Jul 2001 15:54:41 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 23 Jul 2001 12:20:31 +0100 From: Dr.Duncan Clark Subject: Re: DNA sequencing review Message-Id: <20010723145441.CDFED17B8F@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk In article <9jgok3$1ul$1@mercury.hgmp.mrc.ac.uk>, the eminent Ant Smith at BIOSCI/MRC Human Genome Mapping Project Resource Centre wrote >Can anyone point me in the direction of a good review or book that gives an >overview of advances in DNA sequencing technologies from the very first methods, >through radioactive manual sequencing to automated sequencing. My particular >interest is to be able to plot increases in run length and accuracy over the >years and correlate this to changes in the technology used. In addition, I would >like to try to find a plot of $ per base cost over time Maybe the ABRF have this sort of info: ABRF home web site at http://www.abrf.org under Research Committees/DNA Sequence. The ABRF DNA Sequencing Research Committee. This URL is 1999 so I hope it is still OK Duncan -- The problem with the gene pool is that there is no lifeguard. Duncan Clark GeneSys Ltd. Tel: +44(0)1252376288 FAX: +44(0)8701640382 --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 15:55:37 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9A99A17BE3 for ; Mon, 23 Jul 2001 15:55:28 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id C692F17BBE for ; Mon, 23 Jul 2001 15:55:20 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 32CBD17B3F; Mon, 23 Jul 2001 15:55:01 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 23 Jul 2001 12:20:31 +0100 From: Dr.Duncan Clark Subject: Re: DNA sequencing review Message-Id: <20010723145501.32CBD17B3F@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk In article <9jgok3$1ul$1@mercury.hgmp.mrc.ac.uk>, the eminent Ant Smith at BIOSCI/MRC Human Genome Mapping Project Resource Centre wrote >Can anyone point me in the direction of a good review or book that gives an >overview of advances in DNA sequencing technologies from the very first methods, >through radioactive manual sequencing to automated sequencing. My particular >interest is to be able to plot increases in run length and accuracy over the >years and correlate this to changes in the technology used. In addition, I would >like to try to find a plot of $ per base cost over time Maybe the ABRF have this sort of info: ABRF home web site at http://www.abrf.org under Research Committees/DNA Sequence. The ABRF DNA Sequencing Research Committee. This URL is 1999 so I hope it is still OK Duncan -- The problem with the gene pool is that there is no lifeguard. Duncan Clark GeneSys Ltd. Tel: +44(0)1252376288 FAX: +44(0)8701640382 --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 16:19:29 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 8997217B82 for ; Mon, 23 Jul 2001 16:19:27 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 1306F17AE1 for ; Mon, 23 Jul 2001 16:19:25 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id D0CD917BEE; Mon, 23 Jul 2001 16:19:15 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 23 Jul 2001 16:13:32 +0100 From: Scottie Adams Subject: Re: Sequencing directly from BACs Message-Id: <20010723151915.D0CD917BEE@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear Tonto. (I've heard the joke :-)) Try http://www.abrf.org/ and look under Research Groups/ DNA sequence. Look for their 200/2001 study on BACs. http://www.abrf.org/ABRF/ResearchCommittees/DSRG/abrf01_bac_poster.pdf You will find a sample protocol which should work fine for you. Scottie At 3:35 AM +0100 7/22/01, TONTO wrote: >Anyone have any tricks for sequencing directly from BAC clones? >Our core lab has never done it and they would like some input before they do >some runs for us. >I believe they are using the big dye terminator kit and a capillary >sequencing machine. I can get more specific info if anyone has had luck >with this. >Thanks, >Tonto >tontogoldstein@hotmail.com > >--- Scottie Adams aka Pamela Scott Adams Manager Molecular Biology Core Facility Trudeau Institute 100 Algonquin Avenue Saranac Lake, NY 12983 Phone: 518-891-3080, Ext. 115 Fax: 518-891-5126 Email: sadams@northnet.org sadams@trudeauinstitute.org http://www.trudeauinstitute.org http://www.northnet.org/mbcf --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 16:23:33 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id C613B17B82 for ; Mon, 23 Jul 2001 16:23:30 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 7028817AB4 for ; Mon, 23 Jul 2001 16:23:22 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 5B48017BAA; Mon, 23 Jul 2001 16:23:15 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 23 Jul 2001 16:21:38 +0100 From: David Cain Subject: newsgroup update Message-Id: <20010723152315.5B48017BAA@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear Sequencers, When the newsgroup was started in 95 I conducted a small survey looking at capacity, development and how the newsgroup could better support people. I was thinking this morning that a lot has changed in the last 6 years and it may be interesting to see what people currently feel in the so called post genomics era. I have put together a very short survey which I have attached to this email. I would be most grateful if you could take 5 minutes to complete it and send it back to me either via email or fax (+44 20 7720 4798). I am particularly interested in ways that you feel the newsgroup could be developed to better support your needs. Thanks in advance. Sequencingly, David Cain administrator biosci.genome.autosequencing +44 7899 703146 --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 23 17:04:04 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 6FC1B17AD7 for ; Mon, 23 Jul 2001 17:04:02 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id E811E17AF7 for ; Mon, 23 Jul 2001 17:03:58 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 1DBEA17AD7; Mon, 23 Jul 2001 17:03:55 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 23 Jul 2001 17:02:54 +0100 From: David Cain Subject: newsgroup survey Message-Id: <20010723160355.1DBEA17AD7@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Dear Sequencers, I seem to be having problems being able to send the survey form out. To remedy this I am attempting to cut and paste it into this email. Copies are also available if you email me at: david.cain4@btinternet.com Sorry for this hassle, and thanks in advance for your input. Sequencingly, David Cain Genomics users survey 2001 Name: Position: Address: Phone number: Email address: Please give a short description of the role you facility plays (e.g. central resource for academic department) How many people are employed in your facility? What applications do you use and how often? On average how many reactions of each type do you process per day? What analysis systems do you use? (i.e. number of instruments, age and type) What percentage of your maximum capacity do you operate at? Do you anticipate needing to increase the capacity of your facility in the next 12 months? Average read length for each type of template? Do you use reagent saving solutions? (if yes which ones) Do you read the internet newsgroups on DNA sequencing? (if yes which ones) What improvements can you suggest for the newsgroups? What bio-informatics software do you use on a regular basis? Please rate the software you use on a scale of 1 to 10 where 1 is the lowest and 10 the highest? What new software features would help you in your analysis? What new features would you like to see developed for new automated systems? What role does automation play in your laboratory? Do you anticipate that automation will play a more or less significant role in the next 12 months? What meetings/conferences do you attend? What are the main benefits from the meetings/conferences you attend? What do you feel will be the next break through technology for DNA analysis? From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 24 09:25:44 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 92DA417C1B for ; Tue, 24 Jul 2001 09:25:42 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9835217C19 for ; Tue, 24 Jul 2001 09:25:40 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id B7D3B17C1B; Tue, 24 Jul 2001 09:25:36 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 24 Jul 2001 00:42:22 +0100 From: SMarsh3807 Subject: Re: Sequencing directly from BACs Message-Id: <20010724082536.B7D3B17C1B@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk We do this routinely in our Core Facility. We follow the protocol that is in the Big Dye protocol book that comes from ABI. We get great results from that. Big Dye Version 2 seemed to work better for us on the BACs. Steve Director DNA Seq Core Facility @Caltech --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 30 09:00:56 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 8A43D17D48 for ; Mon, 30 Jul 2001 09:00:54 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 87E7B17E1C for ; Mon, 30 Jul 2001 09:00:52 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 9261117D48; Mon, 30 Jul 2001 09:00:47 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 30 Jul 2001 00:52:22 +0100 From: Arthur Mangos Subject: 3700 Wizard problems Message-Id: <20010730080047.9261117D48@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk I am experiencing problems with the wizard software functions on the 3700. When changing arrays the Wizard program didn't complete the full operational function , no acid wash occured. Has any other user experienced this problem when using these Software problems. Any feedback would be appreciated. Arthur Mangos Molecular Patholgy I.M.V.S ADELAIDE SOUTH AUSTRALIA --- From owner-autoseq@hgmp.mrc.ac.uk Mon Jul 30 20:31:21 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 19B3E17AD9 for ; Mon, 30 Jul 2001 20:31:19 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id D48BF17ADF for ; Mon, 30 Jul 2001 20:31:16 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 2D70F17AD9; Mon, 30 Jul 2001 20:31:12 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 30 Jul 2001 15:28:42 +0100 From: Phillip San Miguel Subject: Re: 3700 Wizard problems Message-Id: <20010730193112.2D70F17AD9@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Yes. But it was due to a pump error during the wizard. First I suggest ignoring the wizard's advice to put a dry filter in. Flush the filter with ddH2O and then install it (still filled with water). This puts less strain on the pump. But your real problem is that the wizard has partially completed. This means that the first part of the wizard is done. Attempting to re-run the wizard will then always result in failure. Here is why. The change array wizard is composed of a couple of different parts. 1. POP6 to water 2. install array Lots of pressure can be built up in the cuvette when there is POP in the capillaries--the POP2water part depends on this. But once the POP has been replaced with water there is little back-pressure. So the POP2water part will always fail to reach the desired pressure and error out. Over and over again. The problem is that if there is POP in the caps you definitely do not want to run install array (which includes an acid wash). So it would be best to start the regenerate array wizard (after a complete shut-down, bring-up of the computer and 3700). Then watch hyper terminal. The first part should take about 70 minutes. You can see all the pressure readings as water is pumped into the cuvette and a back-pressure of 120 PSI (1200 units in the hyperterminal window, the pressure readings are in square brackets: [600] [590] [600] ) is attempted. The wizard will repeatedly attempt to reach this back-pressure, then error out. With water the back-pressure is much lower, it will not reach [1200]. Most of the time the wizard will fail to report the error to the 3700 correctly, so the machine will just sit there. But if you see the attempts to pressurize the cuvette failing over and over again. This is irritating, but it does mean that there should be no polymer in your caps, so it is safe to go onto the next part. Shut everything down. Then bring it back up. This time run the "install capillary" choice in the wizard. Don't take your caps out--or you will need to do a new spatial. Just ignore the instructions that relate to putting the array in (you have already have an array.) If you rinse your filter and fill it with ddH2O before installing it, less chance you will overload your pump. So it should go smoothly. If it doesn't work, you might need to open a service call. Phillip SanMiguel Purdue Genomics Core Facility Arthur Mangos wrote: > I am experiencing problems with the wizard software functions on the 3700. > When changing arrays the Wizard program didn't complete the full operational > function , no acid wash occured. > Has any other user experienced this problem when using these Software > problems. > Any feedback would be appreciated. > Arthur Mangos > Molecular Patholgy > I.M.V.S > ADELAIDE > SOUTH AUSTRALIA > > --- --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 08:58:41 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 14D6817A43 for ; Tue, 31 Jul 2001 08:58:40 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 00BE517A58 for ; Tue, 31 Jul 2001 08:58:37 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 291FE17A43; Tue, 31 Jul 2001 08:58:33 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 31 Jul 2001 03:15:00 +0100 From: SMarsh3807 Subject: Re: 3700 Wizard problems Message-Id: <20010731075833.291FE17A43@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk >When changing arrays the Wizard program didn't complete the full operational >function , no acid wash occured. >Has any other user experienced this problem when using these Software >problems. We see this frequently as well. The best thing to do is reboot the 3700 before you start the wizard. Ours usually just kinda stops in the middle, or errors out. Rebooting the 3700 usually works for me. Steve Marsh Director DNA Seq Core Facility @ Caltech --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 13:57:36 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 1C4AA17A89 for ; Tue, 31 Jul 2001 13:57:35 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id A625917A8B for ; Tue, 31 Jul 2001 13:57:28 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 1072A17A89; Tue, 31 Jul 2001 13:57:24 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: Tue, 31 Jul 2001 11:13:55 +0100 From: Hans Sluiman Subject: Sequence Navigator problem Message-Id: <20010731125724.1072A17A89@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Is anyone running Sequence Navigator under Mac OS 9? After upgrading the Mac from OS 8.1 to 9.1 we are no longer able to display electropherograms that were edited previously: "system error 9141 using file xxxxx on volume [HD name]." Assuming this was caused by SeqNav's dislike of the new Mac OS hard disk formatting standard (HFS+) I reinitialized the HD as HFS while keeping OS 9.1. However now we are getting a system error -5016. Since 9.1 is the most robust Mac OS that I know of I'd hate to go back to 8.1 unless absolutely necessary. Any ideas/solutions? Thanks Hans Sluiman -- Dr Hans J Sluiman Scientific, Technical and IT Services Department Royal Botanic Garden 20A Inverleith Row Edinburgh EH3 5LR (Scotland, United Kingdom) +44 (0)131 2482840 (tel.) +44 (0)131 2482901 (fax) From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 14:50:24 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 370B217AA6 for ; Tue, 31 Jul 2001 14:50:20 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9F7E517A95 for ; Tue, 31 Jul 2001 14:50:00 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 8331717A93; Tue, 31 Jul 2001 14:49:51 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 31 Jul 2001 14:45:56 +0100 From: Phillip San Miguel Subject: Re: 3700 Wizard problems Message-Id: <20010731134951.8331717A93@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk SMarsh3807 wrote: > >When changing arrays the Wizard program didn't complete the full operational > >function , no acid wash occured. > >Has any other user experienced this problem when using these Software > >problems. > > We see this frequently as well. The best thing to do is reboot the 3700 before > you start the wizard. Ours usually just kinda stops in the middle, or errors > out. Rebooting the 3700 usually works for me. > Steve Marsh > Director DNA Seq Core Facility @ Caltech This could be similar to the polymer change wizard erroring out immediately after starting it. I've notice this almost always happens if it is run with Data Extractor open. If I close Data Extractor and Data Collection. Then reopen Data Collection and run the polymer change wizard, I never have this error. My problems with regenerations seemed to be caused by a bum polymer pump my machine was shipped with. We nursed it through about 1000 runs before our service engineer took mercy on us and replaced it. Since then, no problems. Also I usually check the hyperterminal log files. I could see we were getting pump overload errors. Phillip SanMiguel Purdue Genomics Core Facility --- From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 16:41:17 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 3DF6317AB6 for ; Tue, 31 Jul 2001 16:41:15 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 07E3117AAC for ; Tue, 31 Jul 2001 16:41:12 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 8B19717AB2; Tue, 31 Jul 2001 16:41:09 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: Tue, 31 Jul 2001 15:04:17 +0100 From: Dr N.I.Leaves Subject: Re: Sequence Navigator problem Message-Id: <20010731154109.8B19717AB2@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk what do ABI say? i dont suppose they suport Navigator in any way any more. nick end ************************************************** Dr N I Leaves Mouse Sequencing MRC HGMP Resource Centre Hinxton Cambridge CB10 1SB tel: 01223 494557 (office) or 01223 494541 (lab) email: nleaves@hgmp.mrc.ac.uk ************************************************** On 31 Jul 2001, Hans Sluiman wrote: > Is anyone running Sequence Navigator under Mac OS 9? After upgrading the > Mac from OS 8.1 to 9.1 we are no longer able to display > electropherograms that were edited previously: "system error 9141 using > file xxxxx on volume [HD name]." Assuming this was caused by SeqNav's > dislike of the new Mac OS hard disk formatting standard (HFS+) I > reinitialized the HD as HFS while keeping OS 9.1. However now we are > getting a system error -5016. > > Since 9.1 is the most robust Mac OS that I know of I'd hate to go back > to 8.1 unless absolutely necessary. > > Any ideas/solutions? > > Thanks > > Hans Sluiman > > > -- > Dr Hans J Sluiman > Scientific, Technical and IT Services Department > Royal Botanic Garden > 20A Inverleith Row > Edinburgh EH3 5LR (Scotland, United Kingdom) > +44 (0)131 2482840 (tel.) > +44 (0)131 2482901 (fax) > > From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 16:42:59 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id E490617AE2 for ; Tue, 31 Jul 2001 16:42:52 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id BC93017ACE for ; Tue, 31 Jul 2001 16:42:40 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 0952A17ADB; Tue, 31 Jul 2001 16:41:50 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: Tue, 31 Jul 2001 15:12:06 +0100 From: Dr N.I.Leaves Subject: Re: 3700 Wizard problems Message-Id: <20010731154150.0952A17ADB@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk I think there are two problems here. Firstly the problem which I think almost everyone is familiar with which is the wizards crashing almost immediately, before they begin really. In our lab this is cured every time by just restarting data collection. I think this is just a bug and is not really important. Second, is the initial problem of Arthurs where the regenerate array wizard has crashed half way through. Phillip's answer seemed very technical and probably accurate. My instinct would be to rerun the wizward and then on failure I would make a Service call to ABI. I'm sure that doesnt help :-( Nick Leaves end ************************************************** Dr N I Leaves Mouse Sequencing MRC HGMP Resource Centre Hinxton Cambridge CB10 1SB tel: 01223 494557 (office) or 01223 494541 (lab) email: nleaves@hgmp.mrc.ac.uk ************************************************** From owner-autoseq@hgmp.mrc.ac.uk Tue Jul 31 21:04:59 2001 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id AB87A17A9D for ; Tue, 31 Jul 2001 21:04:57 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 9800317AF6 for ; Tue, 31 Jul 2001 21:04:55 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 60001) id 50B5817A9D; Tue, 31 Jul 2001 21:04:52 +0100 (BST) To: autoseq@net.bio.net Newsgroups: bionet.genome.autosequencing Date: 31 Jul 2001 19:23:06 +0100 From: SMarsh3807 Subject: Re: Sequence Navigator problem Message-Id: <20010731200452.50B5817A9D@mercury.hgmp.mrc.ac.uk> Sender: owner-autoseq@hgmp.mrc.ac.uk Precedence: bulk Try formatting the disk in HFS mode not HFS+. Also be very careful running Norton utilities on that Hard Drive now. there have been a lot of problems running Norton Speed Disk on drives that were reformatted with HFS+. Steve MArsh ---