Next Generation Sequencing
Introduction
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The MRC WIMM Next Generation Sequencing Facility is located in the laboratories in the basement of the Women’s Centre. Onsite are two NextSeq 500 instruments and a MiSeq instrument. The Facility only processes pre-made libraries. The instruments are covered by a Silver-Service contract, which means that if a run fails for a machine fault Illumina will replace the kit. |
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Using the Facility
General protocol for requesting a time slot on an NGS instrument:
1 - Prepare a library of DNA/RNA using whichever method is compatible with the Illumina platform (e.g Fluidigm Access Array, NEBNext, Nextera, TruSeq etc)
2 - QC the library – and try to get either a 2nM or 4nM pool
3 - Use the Agendo system to select a free time slot and email Tim Rostron to request the booking.
Information required:
- PO Number
- Date
- Instrument
- Kit
- Output
- Cycles required: Read1= i7= i5= Read2=
- CCB upload required: Yes/No If yes, provide your CCB username
- BaseSpace required: Yes/No If yes, please provide username/password (strictly confidential)
- Library concentration:
- Special instructions: e.g. custom recipe or custom primers
4 - Make a sample sheet using a suitable template (templates available on request)
5 - Use R12 to generate a PO before the day of the run.
Sample/custom primers/kit components should be brought to the sequencing fridge at 10 am on the day of the run.
Kits:
We do not offer a library preparation service or keep kits in stock, so users must do their own library preparation and purchase their own kits.
If you are a member of the University of Oxford, you will get a 20% discount from Illumina. Purchasing of kits can be done using R12 (- remember to buy the kits in USD).
Purchase Orders:
The WIMM Sequencing Facility will require a Purchase Order (PO) to be raised in R12 for all MiSeq and NextSeq work. The supplier in R12 is 'Weatherall Institute of Molecular Medicine'.
Current prices can be found on our intranet.
Agendo Account:
If you are already using the Agendo system for FACS/microscopy bookings, you do not need to register again. If you don't have an Agendo account, go to the Agendo booking system, click [New User] and fill in the details. The Resource is either Illumina MiSeq (Sequencing Facility) or Illumina NextSeq (Sequencing Facility).
Results:
Results are available either from BaseSpace if required or by upload to the CCB server.
Having a BaseSpace account will enable you to monitor runs from the MiSeq and NextSeq500 within any browser or from the BaseSpace App. Accounts can be created from the Illumina website.
If you intend to use the CCB facilities for analysing your data, you must have a CCB account and it is prudent to enlist the assistance of a bioinformatician first, to ensure that you have the right post-run pipeline in place for the data you are generating. If you need a CCB upload, NGS data will be transferred to a holding folder and you will be sent an email when the upload is complete.
Further Information:
If you require further information or csv templates, please contact Tim Rostron