DNA Sequencing and Fragment Analysis Service

Sample submission

The Facility installed a new ABI 3730 state-of-the-art capillary Genetic Analyser on March 3, 2003. The Facility has a long history of DNA seqeuncing with the installation in 1988 of an ABI 370 with 16 lane capacity (initially run by an HP Vectra computer). This was later upgraded to an ABI 373. In 1996 the Facility acquired two ABI 377 DNA Sequencers which were subsequently upgraded from 48 to 96 lane capability.

The new 3730 has 48 capillaries (with an option to upgrade to 96) and a run-time of 2h using 50 cm capillaries. This means we can run a 96-well plate in 4h and finally gives us the capability for high throughput project sequencing. The new technology in conjunction with BigDye 3.1 enables longer read lengths to be obtained, an improvement in quality and reduced cost (less BigDye usage and cheaper per reaction costs).

We aim for a turnaround time of 1-3 working days for the standard service (user prep) and 5 working days for the full service (core prep). Internal controls are included in all sequencing runs. Our pGEM controls routinely produce over 1000 base pairs of data with a Quality score >20.

Remember the quality of your sequencing is determined by the quality and quantity of DNA template and primer that you use. Clean-up of sequencing reactions is now critical as large dye blobs will saturate the signal and salt will prevent correct electrokinetic injection of samples.

Please read our guidelines for sample submission and quantities of template/primer to use/submit for your reactions. We recommend that you print out a copy of the relevant protocol for sequencing with BigDye and read these carefully before commencing DNA sequencing. Read the section on Template Preparation as it contains a lot of useful information including info on estimating the concentration of template. Please check out the trouble-shooting pages if you are having difficulty obtaining good sequencing results.

We prefer to perform sequencing reactions at the Facility to reduce variability in sequencing results.