|To create an account on our iLabs site, download these instructions. Contact Mark Farrell with any questions or concerns when creating a new account.|
- Standard Reaction/Data Collection: samples should be submitted in the following form:
- For plasmid DNA <12kb - provide 20 µL of DNA at a concentration of 40 ng/µL in water. For PCR products 300-600bp - provide 20 µL of DNA at a concentration of 10 ng/µL in water.
- For a custom primer, please provide 10 uL at a 10 uM concentration. The primer should be designed to have a Tm as close to 55C as possible. When sequencing multiple samples with the same primer, please provide 10 uL for the first template+primer combination, and 2 uL for each combination after that. For example, when submitting 10 samples to be sequenced with one primer, please submit 10uL for the first sample + 2uL x 9 remaining samples for a total of 28 uL. This being in contrast to 10 uL x 10 samples for a total of 100uL.
- When submitting the same sample to be sequenced with multiple primers, please submit 20uL volume of template for the first template-primer combination and 5 uL volume of template for each template-primer combination after that.
- For template/primer mix please provide a 20ul volume of template at 40ng/ul for plasmids or 10ng/ul for PCR products and primer at 1uM final concentration.
- For PCR fragment DNA- amplicons must be purified/treated prior to DNA sequencing. Please inquire for details and/or protocols. If you would like to submit PCR reaction, we can treat/prepare your sample for sequencing by purification or ExoSap treatment. Please inquire about cost and sample submission.
- The following standard primers are provided: M13F(-20), M13R,T7, T7 Term, SP6, T3, CMV-F, PGEX-F, PGEX-R, BGH-R, Lucif Ctrm Rev, GFP-R, pNTR1A-F, pNTR1A-R, PQE30-F, PQE-Tri-F, PQE-Tri-R, E-GFP-R, and V5 Ctrm Rev.
- DNA samples that are submitted for sequencing must be accurately quantified and prepared. Samples submitted with inaccurate concentrations and/or with contaminants present will result in poor sequence results or a failed reaction.
- After processing the results will be available to you from the iLabs site. Please login and click on “View My Requests” tab. Click on the triangle corresponding to your order to expand it out. Your results will be attached to your order in the form of a .zip file under “Attachments and URLs”.
- The sequence will be repeated at no extra cost if there is an operator error or machine malfunction. Please provide accurately quantified, high quality DNA for successful results.
- Automated sequencing is very sensitive to template and primer preparation.
- It is essential that the templates be free from ethanol, salts and other contaminants, as these greatly affect the quality of the sequencing results.
NOTE: WE WILL NOT REPEAT SAMPLES AT NO CHARGE IF THE DNA IS OF BAD QUALITY OR INACCURATE CONCENTRATION.
Here are a couple programs available to view chromatogram files (.ab1):
Chromas Lite – from Technelysium http://technelysium.com.au/?page_id=13
Finch TV – free download from Geospiza http://www.geospiza.com/Products/finchtv.shtml
Chromatogram Explorer – from DNAbaser http://www.dnabaser.com/download/chromatogram-explorer/
Human Identity Testing, Cell Line Authentication, and Mycoplasma Testing:
- Please provide isolated/purified genomic DNA in a minimum volume of 15ul and a minimum concentration of 30ng/ul.
- For ID and CLA testing only- a cell pellet can be submitted and we can perform the isolation for you. Standard DNA isolation rates apply.
Barbara Davis Center
Anschutz Medical Campus
1775 Aurora Court, 4th floor, Room 4207
Molecular Biology contact personnel: Mark Farrell, Core Manager