DNA Sequencing Service

DNA sequencing is performed on the Applied Biosystems Model 3100, a capillary based electrophoresis system. The system uses a fluorescent dye terminator chemistry. For details on the features of the instrument and chemistry see the Applied BioSystems website. We have two of these instruments, each is capable of sequencing 16 samples in 2.5 hr. up to 196 samples (about 120Kb) per 30 hr. run. There are two methods of submitting your sample for DNA sequencing. You may do the reactions and subsequent purification of the extension products yourself; please see these instructions. If you choose not to do the sequencing reactions or the purification the Core lab will do the reactions at additional cost, provide us with template and primer or just template if a standard priming site is available on your template according to the following guide lines. We will perform the sequencing reactions and cleanup of the extension products. See the current order form for pricing.

If you have sufficient non-problematic template a good primer and clean extension products, it is not unusual to get 600-700 bases of unambiguous data. Once you develop some proficiency at editing the computer called sequence you can often get another 100 bases. The standard we run on every gel typically sequences to 97% accuracy at 700 bases, unedited.

When the sequencing is completed you will receive your data by e-mail. Each sample will have two files associated with it . The ABI Prism 3100 DNA Sequencer operates under Windows NT therefore the data files are in windows format and will need to be converted to be viewed on the MAC. The freeware conversion program is available from our folder on the Dartmouth public server or from Applied BioSystems web site. (If you are a PC user, you can download software from Chromas to view and edit your data.) One file will show the electropherogram for your sample. This file can be edited to correct problems with the computer called sequence or EditView1.0.1. You will also receive a file with a .seq extension. This file is a text file with raw sequence called by the computer. This file can be viewed using Microsoft Word.

Order forms are available at the laboratory, 201 Vail, by BlitzMail, or from the Molecular Biology Core folder on the Dartmouth Public server. Please fill out the order form according to the instructions given. Completed order forms and your sample can be dropped off at any time at the laboratory, 201 Vail, or at the Borwell pick up site Room 716E. Data from samples submitted before 3:00 at 201 Vail before 11:30 at 716E Borwell will usually be reported to you the next day.

The cost of a DNA sequence is $6.50 per lane for completed reactions. $17.50 per sample when we perform the sequencing reaction and purification of extension products. See the sample sheet for a complete price list.

Turn around time is usually 24 hours or less for completed reactions. However this is highly dependent on the number of samples in the queue. If we do the chemistry the turn around is 72 hours or less depending on the time of day the samples arrive at the laboratory. For samples that arrive at the laboratory by noon, under the current sample load (3/03), we can usually get your data out the following AM. If your samples are at the lab before before 3:00 PM they will be included in a later run but may not finish by the AM. The data will be reported out as the samples finish during the following day.