Accelerated discovery of functional non-coding genomic variation using single molecule real-time (SMRT) sequencing.

The last decade has seen tremendous advances in our understanding about the role of genetic variation in health and disease, driven largely by developments in technology for sequencing the genome. Genes involved in a spectrum of debilitating disorders and health-related traits have been identified, including many by researchers at the University of Exeter Medical School, and these discoveries are starting to directly impact upon clinical practice. Despite this success, however, many regions of the genome remain hidden to contemporary first- and second-generation sequencing approaches. These include highly complex stretches of repetitive DNA and regions harbouring other types of genetic variation such as deletions and duplications. Furthermore, because the sequencing reads generated by current approaches are relatively short, they cannot be used to examine the long-distance relationships between more distal variants, nor explore the extent to which different genes are expressed in different versions in health and disease. Finally, most focus in disease genetics has been on regions of the genome that actually encode specific genes, ignoring the role of regulatory variation occurring outside of these regions. Of note, current sequencing approaches cannot be used to directly detect the presence of non-sequence-based epigenetic modifications and explore their role in disease. In this application we are requesting funds to support the acquisition of "third generation" sequencing technology, which will enable us to fully characterise genomic complexity. We are also requesting funds to support the high-performance computing infrastructure needed to store and analyse this complex data. Our aim is to advance genomics research into diabetes, neurological and other medical conditions by defining functional genetic and epigenetic variation in non-coding regions of the human genome.

We are requesting funds to develop our genomic sequencing infrastructure, primarily through the acquisition of a Pacific Biosciences single molecule real-time (SMRT) sequencing system, making us one of only a handful of institutes in Europe to have such capability. This system will enable us to assay, identify and analyse regions of the genome hidden from current methods for gene discovery, and facilitate research into the functional mechanisms involved in disease. The Pacific Biosciences RS II SMRT Sequencing System provides the highest consensus accuracy and longest read lengths of any sequencing technology; current average read-lengths are >8.5kb (with a maximum of ~50kb). SMRT sequencing will facilitate advances in disease characterisation, stratification and diagnosis via its unique utility for de novo assembly across highly complex regions of the genome, targeted re-sequencing, long-range phasing of genetic variation, and epigenomic analysis via the direct allele-specific detection of modified DNA bases. We will also develop our High-Performance Computing/Informatics Infrastructure to enable the storage, analysis, alignment, and assembly of SMRT sequencing data. This infrastructure investment will enable the world-leading genetic and epigenetic research groups at the University of Exeter Medical School to further develop their innovative program of clinical genomic research in human disease, with a specific focus on diabetes and neurological phenotypes.

In addition to other scientists (especially other and research groups investigating the role of genetics in diabetes and neurological diseases) the data generated using the new sequencing infrastructure has the potential to impact on a number of other beneficiaries. These include patients, the pharmaceutical industry, health service providers, and academic groups investigating the causes of other disease phenotypes and functional genomics. Existing genetics research by Exeter groups has already made significant impact in terms of understanding the causes of disease and the mechanisms behind disease progression (see description in the Case for Support), with tangible patient benefit. For example, Professor Hattersley's group discovered that patients with neonatal diabetes caused by K-ATP channel mutations demonstrate a remarkable improvement in glycaemic control when sulphonylurea tablets replace insulin injections. Quality of life has also been improved for many patients with maturity onset diabetes of the young (MODY) who have either stopped treatment or transferred from insulin to sulphonylureas as a consequence of their genetic diagnosis. The ability of SMRT sequencing to interrogate regions of the genome hidden from second generation sequencing, in addition to epigenetic and transcriptomic variation, will identify additional pathways to disease that we hope to have similar impact on clinical diagnosis, prognosis, and treatment.