Next Generation Sequencing Facility
- Overview: The NGS facility is owned and run by the University of Leeds. We aim to provide a flexible sequencing service in which cutting edge methodologies can be rapidly developed and provided, as a service to both internal and external users.
The facility is currently equipped with Illumina HiSeq 3000, NextSeq and two MiSeq sequencers as well as a PacBio Sequel and ONT Minions, and we can discuss which of these may meet your needs for sequencing of DNA or RNA samples. Depending on requirements, we can sequence user-prepared libraries or provide an extensive library preparation service. The facility has a proven track record in the effective use of exome, polyA selected, rRNA depleted and miRNA RNA-Seq and Chip-Seq methodologies, and we are continuously striving to broaden our skill base.
As our team works closely with members of the NHS diagnostic labs and University research staff, we are able to generate data with a full audit trail to NHS diagnostic standards while simultaneously proving a flexible environment that allows us to rapidly respond to new developments in your research field.
- HiSeq 3000: This machine can generate >1000 Gb of data per run and is also suitable for a wide range of sequencing application.
- NextSeq: This machine can sequence >400 million inserts per run and is also suitable for a wide range of sequencing application.
- MiSeq: With a run time of as little as 24 hours, this machine can generate up to 5.1 Gb of data when performing a 150-bp paired reads run.
- PacBio Sequel: The PacBio is able to sequence a wide range of templates to generate high quality reads approximately 3,000 bp in length
- ONT Minion: The Minion can routinely generate read sequence tens and even hundreds of kilo-bases in length. While the base calling quality is low, the read length is ideal for the detection of structural variants and de novo assemble
The NGS facility has access to all the equipment needed to create libraries for your sequencing projects. For many applications, we can offer to prepare libraries for you. Alternatively, it may be more appropriate for users to generate their own libraries, and in this case, we can discuss arrangements for access to our facilities. Read more
We have considerable expertise in analysis of large-scale sequence data, either using standard research techniques or by developing new analysis methodologies in partnership with investigators. We will support you in whatever way we are able. We can also offer assistance in setting up your own personal analysis platform on the University's HPC clusters (ARC2, ARC3 and ARC4 when it's operational). Currently, we routinely perform RNA Seq analysis, exome based variant detection and annotation, de novo assembly of genomes (bacterial and viral) and transcriptomes, 16S/18S meta-genomics analysis and variant calling for bacterial whole genome sequencing Read more
To date we have developed a range of bioinformatics programs that facilitate various tasks across a wide range of medical and biological disciplines. Currently available software applications can be found here.
We are experienced in a wide range of library production techniques and aim to add more methodologies to our repertoire as time progresses, allowing you access to the most up to date methods in your field. If you have any questions or suggestions please contact us.
We are able to offer a range of standard library preparation, but are also willing to help you create your own. Whether you require help troubleshooting, tips on best practice, or just an aliquot of a library preparation kit, we are prepared to help.
Below is a list of commonly used library preparation kits with possible suppliers if you wish to buy your own.
- For DNA Library Preparation, for applications such as whole genome sequencing, amplicon sequencing and copy number variation (CNV), we recommend the
NEBNext Ultra DNA Library Prep Kit for Illumina with
NEBNext Multiplex Oligos for Illumina (Index Primers Set 1 or Index Primers Set 2).
- For mRNA Seq Library Preparation, we recommend
TruSeq Stranded mRNA kits with the appropriate indexes from Illumina sequencers, which gives strand directional information on transcripts over ~ 250 bp with a polyA tail.
- For RNA Seq (mRNA and lncRNA) Library Preparation, we recommend
TruSeq Stranded Total RNA Human/Mouse/Rat or TruSeq Stranded Total RNA Gold kits with the required rRNA, mitochondrial and/or chloroplast depletions reagent and indexes from Illumina sequencers. These give data on any transcript over ~280 bp that isn't removed by the depletion reagent.
- For miRNA Library Preparation, we recommend
NEBNext Multiplex Small RNA Library Prep kit or QIAseq miRNA ibrary Prep kit for Illumina sequencers (with the required index kit).
- For Exome analysis, we recommend the Agilent SureSelect XT system. There are two elements to this;
SureSelect XT Reagent kit and SureSelect XT Human All Exon V5.
Please Note: All these kits contain the basic reagents required for library preparation, but ALL require further components to be purchased separately.
We recommend purchasing NEB kits from Science Warehouse and Illumina Kits directly from Illumina (by creating an online account at illumina.com).
If you would prefer to have your libraries made in house for you, there is a fixed charge per library depending on type and number of samples. A price list can be found here. When viewed via the University of Leeds intranet the price will not include VAT, but will include VAT when viewed off campus or via a non-UoL network such as on a mobile phone connected via eduroam.
If the payment comes from a non-University of Leeds account VAT will be charged.
We very happy to discuss your requirements and answers questions and queries (emails here).
Our prices are comparable to those to those of other academic research facilities and typically much cheaper than commercial service providers. Read more
We have a HiSeq 3000 sequencer which typically generates 150 bp paired end sequence data. As soon as your Illumina-compatible sequencing libraries are prepared, please contact us to reserve your place in our sequencing queue. Samples are run on a first-come, first-served basis. The run time for a standard 2x150-bp run takes approximately 3 days, plus a daya to base call the data. Since the HiSeq processes 8 pools in a single run, it is operated once every 2 to 4 weeks. The output is supplied as standard gzip compressed Fastq files.
Illumina certified HiSeq service provider
The NGS Facility is now an Illumina certified sequencing provider after demonstrating an in-depth knowledge of sequencing, library production and data analysis resulting in the production of high sequence data.
Learn more here.
As well as the HiSeq 3000, we have a NextSeq that processes one pool per run allowing it to be for more flexible than the HiSeq, although at a high per-base cost. we offer a range of NextSeq run type such as 75 bp single and paired end runs with the paired end runs available in high (over 400 million inserts) or mid (over 130 million inserts) output options. Typically there is only a short queue to use this instrument with 1x75 bp sequencing processed within 10 working days (including how QC steps for each pool). Other run types may take longer to process depending on how quickly we can get the reagents from Illumina. The output is supplied as standard gzip compressed Fastq files.
Learn more here.
Illumina MiSeq runs produces paired-end data with read lengths of 150 bp, 250 bp or 300 bp. The run time for a standard 2x150bp run is 24 hours with the 300 bp reads taking just under two days. Typically there is only a short queue to use this instrument with 250 bp runs processed within 10 working days (including how QC steps for each pool). Other run types may take longer to process depending on how quickly we can get the reagents from Illumina. The output is supplied as standard gzip compressed Fastq files.
Learn more here.
We have a PacBio Sequel sequencer which typically generates several hundred thousand ~3 kbp circular consensus sequences (CCS) of high quality sequence approaching that of an Illumina short read sequencer The output is supplied as standard zip compressed Fastq, fasta or unaligned BAM files. We have had several success de novo assembling bacterial genomes consisting of a single contig per chromosome. We have also developed pipelines to sequence and detect variants in very repetitive GC rich amplicons, which can't be easily sequenced with Sangar or Illumina sequencing
The NGS service is based on the University of Leeds's St James's University Hospital campus. For all enquires please send an email to Dr Ian Carr.
University of Leeds
- Ian Carr: Academic Lead
- Morag Raynor: Senior Technical Specialist
- Ummey Hany: Technical Specialist
Next Generation Sequencing Facility email
- Dr Ian Carr: Academic lead