DNA Sequence Laboratory

The DNA Sequence Laboratory provides services for the sequence determination of DNA, as well as applications in fluorescent fragment analysis. DNA sequence is determined following cycle sequencing with thermostable polymerases and fluorescently labeled dideoxy terminators. State of the art, capillary based, automated DNA sequencer platforms are utilized to analyze sequence derived from various sequencing protocols and template types. Fluorescently labeled fragment analyses for genetic/population studies are carried out utilizing Genescan (TM) software. Information on setting up an account and submitting samples can be found in the "First Time Users"  and "Pricelist and Submission Information" documents at the DNA files page. Data is recovered over the internet as described in "Computer Information" at the DNA files page.

All data and reagents distributed by the University of Wisconsin Biotechnology Center DNA Sequence Facility are intended for research purposes only.  They are not intended or certified for diagnostic or clinical use.

Sequencing Options

A number of service options for DNA sequence determination are available through our facility. The most economical service (formerly "Plan I") involves the user setting up their own reactions, then bringing the cycle sequence products to our facility. We can provide the sequencing reagent at a very low rate thanks to a favorable University quote, and go over the protocols with you necessary to carry out the reactions in your laboratory (a PCR machine is the only required equipment). Various sequencing protocols that have worked for different groups can be found in the document "Cycle Sequencing Protocols" at the DNA files page. At that same link is a document describing the particular protocols we use in the facility, called "Facility Procedures." We also can provide positive controls to help you get started (as described in the document "Available Controls" at the DNA files page, or troubleshoot when results are not satisfactory. Numerous price options for submitting your sequenced reactions are possible depending on the number and format of your samples, check the document "Pricelist and Submission Information " at the DNA files page for a description.

We also can provide expert service in sequencing if the user wishes simply to provide the template DNA (formerly "Plan II"). We can sequence double strand plasmid, single strand phage, PCR products, BACs, cosmids, and related vectors. Common vector primers are provided (e.g., SP6, T7 M13 F and R, etc.-check availability). Every effort is made to provide at least 600 bp of excellent sequence (Q value >20), though template composition and purity are crucial determinants of quality. Strategies are available to sequence difficult GC rich templates or homopolymer regions, user should indicate if such areas are present. Other information regarding dropped off DNA samples can be found in the documents titled "First Time Users" and "Pricelist and Submission Information"at the DNA files page. Certain discount sequencing options are also available and described in the above documents.

The DNA sequencing facility can also carry out larger sequencing projects involving editing and

assembly (formerly Plan III). The maximum size of the sequence we can generate is approximately 20 kb. Price is based on the degree of confidence required (i.e., single strand vs. double strand, one vs. multiple passes); check the price list or call for more information.

Finally, a number of different options are available for the processing of 96 well plates. Please consult the price list to see what we can do for you if samples are in, or could be generated, using this format.

The UWBC DNA Sequence facility can also work with researchers to carry out services that may not be listed in our current rate schedule. We can generate custom charges based on our estimates of what the work will cost. If you don't see something you want us to do, please inquire whether we can carry out this service.

Fluorescent Fragment Analysis

For this service, the user generates fluorescently labeled PCR fragments (typically through use of fluorescently labeled primers, available through our DNA Synthesis facility, or through incorporation of fluorescent nucleotides) that are resolved and accurately sized on our automated sequencers using the Genescan software. Suitable for microsatellites, AFLP, RAPDs, and related procedures. Price depends on the number of samples. For more information, consult the "Fragment Analysis" document at the DNA files page.

Policy Statement

Every effort is made to provide our users with the best data in a timely fashion for the advancement of their research. Turnaround time is typically one working day. If you haven't seen your data on the server by three working days, contact us. Samples that have failed due to gel or capillary problems will of course be rerun free of charge. Usually such problems have a characteristic appearance, so contact us to see if this is where your difficulty might be. Samples are kept seven days, so you must let us know within that time frame if your samples need to be rerun. Our facility handles 4-5,000 samples/week, and despite our best efforts we occasionally make mistakes. If you get data that is completely puzzling (e.g., sequence from an organism that's not even in the same phylum as your model system) please contact us as soon as possible and we can re-run the samples. As above, we need to know within seven days so we can rerun the same samples you brought over. Sometimes we can even trace the problem and give you the correct sequences right away. If the questionable samples are gone, the initial analysis and reanalysis will be done free of charge, and a reasonable attempt made to compensate for reagents wasted.