The AEGRC has been operating a small in-house sequencing service for UQ researchers since 2007. We offer high quality, fast, customized services, all our machines are highly maintained and controls are included in each batch of sequencing. We also provide customised solutions such as different types of sequencing dyes for particularly difficult DNA templated eg. hairpin or highly repetitive regions.


Result from pGEM-3Zf(+) internal control run with every batch of sequencing

 

The sequencing services offered by the AEGRC include the following:

Both the Sanger DNA sequencing service and the Fragment analysis service are performed on our capillary electrophoresis machines. The AEGRC uses an Applied Biosystems 3730xl Genetic Analyser, and processes both BDT v3.1 and BDT v1.1 sequencing reactions.

Our Sanger Sequencing and our Fragment Analysis service run each business day (excluding public holidays) at 12 noon. Samples submitted for our Clean-up and Separation only service will be reported the following business day if submitted prior to 12 noon. Samples requiring our Full Sequencing service take an additional day, and will be reported in 2 business days following submission prior to 12 noon.

If you are interested in our sequencing or fragment service, please email uqaegrc@uq.edu.au for a submission form and a RISQ form. The RISQ form will need to be completed and returned prior to routine sample submission.

 

Sanger DNA sequencing 

We offer two types of Sanger sequencing services:

"Full sequencing service" – you provide the template and primer and we do the sequencing reaction, clean it up and separate on our capillary electrophoresis machines.

For the full sequencing service we do require the DNA and primers to be pre-mixed. We ask for the DNA, primers and water (if necessary to make up the volume) to be submitted as a total volume of 15.5ul in a 1.5ml tube. The primer concentration is 3.2pmol in a 20ul reaction. The DNA concentration depends on the actual sample.

"Clean-up and separation only" – you do the sequencing reaction, we clean it up and separate it on our capillary electrophoresis machines. 

Our sequencing reaction setup guide has suggested reaction volumes and cycling conditions and can be provided on request. Most people start with using 1ul of Big Dye and move to 0.5ul or less once sequencing is optimised.

We also sell the BigDye sequencing reagents in small aliquots (so customers do not have to buy whole expensive kits) if you are interested in doing the reaction yourself. 

Results

All sequencing files are saved onto a special shared drive set up for clients. Only people that have approved access for each group will have access to that individual group’s results folder. Results will be made available on the shared drive for a 1 month period, however, the AEGRC maintains an archive of all sequencing results and can reissue past results on request.

 

Fragment analysis 

We offer a Fragment analysis service for capillary separation and size determination of fluorescent labelled PCR products. For this service you perform the PCR and provide us with the PCR product for running on the DNA analyser. Dyes that can be used include 6-FAM, VIC, NED or PET.

Results

All fragment FSA files are saved onto a special shared drive set up for our clients. Only people that have approved access for each group will have access to that individual group’s results folder. Results will be made available on the shared drive for a 1 month period, however, the AEGRC maintains an archive of all fragment analysis results and can reissue past results on request.

To analyse your FSA files you can use Peak Scanner (free online software).

Sample submission

Samples can be submitted in person to the AEGRC.

Sample Submission Times: 8:00am to 4.30pm (excluding weekends and public/University holidays).

AEGRC Location: Level 7, Chemistry Building (#68), University of Queensland, St Lucia.

Alternatively, samples can be submitted via the internal mail or sent via Australia Post.

Sequencing Samples

Please submit samples for clean-up and separation in 0.2ml strip tubes. Samples should be wrapped in alfoil to protect them from the light. Full sequencing samples can be submitted in 1.5ml tubes. A submission form must also be emailed to uqaegrc@uq.edu.au before we can process the samples.

Submitting large batches of samples?

For large batches of samples (greater than 32 samples), please submit samples in 96-well plates. For clean-up and separation only samples, please leave one well empty for our internal control. For full sequencing 2 internal controls are used (1 for the full sequencing reaction and 1 for the clean-up and separation reaction) so 2 wells will need to be left empty on these plates.

You will receive a 10% discount for FULL plate submissions – 95 samples for our clean and separation service and 94 samples for our full sequencing service.

How should I label my samples?

Please use a simple and short labelling system for your samples. The labelling on the sample tubes will need to match up with the sample name on the submission form, so please make sure you choose a sample name that can fit on your sample tube. We suggest using the system of your initials, followed by the sample number, eg. AB1, AB2, AB3 etc. We will also add a unique batch number to your sequencing name, so if you are re-using sample names you will be able to differentiate the result.

Please label tubes on the lid and the side with a blue or black permanent marker. For 96-well plates, please write your name and plate number (if submitting more than 1 plate) on the front of the plate.

Our genetic analyser does not accept spaces in sample names, or characters other than numbers and letters, so please make sure you do not include these in your sample names or we will need to alter the name.

The submission sheet should be filled out from top to bottom, left to right eg. A1, B1, C1, D1, E1, F1, G1, H1 then A2, B2, C2 etc. Please make sure the position of your samples match up with your submission sheet. If any tube or well is left empty, please leave this position empty on the submission form.

Fragment samples

Please submit samples for fragment analysis in strip tubes or a 96-well plate. Please submit approx. 10ul for each sample. Samples should be wrapped in alfoil to be protected from the light. A submission form must also be emailed to uqaegrc@uq.edu.au before we can process the samples.

 

Financial information

The AEGRC will send you a RISQ Form (Request for Internal Services Quote) for sequencing and fragment analysis services that will have to be signed by the Research Group Leader, usually the “Budget Holder”, and signed by the School/Institute Finance Officer, usually the “Financial Delegate”. UQ Researchers can use a RISQ Form to set an Upper Limit of expenditure on sequencing over a 12 month period. The RISQ Form does NOT initiate any transfer of funds. It is a Quote, where the AEGRC specifies services and prices and Researches agree to pay for the services requested.

Sequencing services are invoiced by the ISC at the start of each month for samples submitted in the previous month.

For more information regarding billing, including cost, please contact the AEGRC at uqaegrc@uq.edu.au