High throughput intermediate scale sequencing for the Midlands

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Since the advent of sequencing, driven by Frederick Sanger, the knowledge of the underlying genetic code of an organism has been a key feature in establishing the mechanisms by which inheritance influences phenotype. The rapid explosion in next generation sequencing technology in the past decade has taken us from huge consortia trying to determine the sequence of a single human genome to individuals being able to sequence their own genomes for close to $1,000. Alongside the sequencing of the genomic DNA it is also possible to sequence the transcriptome (genes expressed in a cell or tissue) or investigate modifications to the bases of DNA that alter the expression of genes. Researchers at the University of Nottingham have used NGS to investigate changes in the copy number of sequences within the genome and even used NGS to investigate how genomes are copied during bacterial replication.Although the UoN has a range of sequencing technologies available, there is a clear gap at two levels of sequencing that are addressed by this proposal. Current technologies available at UoN allow sequencing at the level of individual microbial genomes or targeted sequencing of small regions of the human genome. These scales preclude the use of NGS to study diverse organisms with larger genomes where many of the techniques developed for studying well-characterised genomes can be readily applied. At the opposite scale, whole genome sequencing (WGS) of small genomes can be of enormous importance when carrying out both targeted and random mutagenesis experiments to engineer new phenotypes (synthetic biology). Here the generation of compensatory or off target mutants is a risk and so routine WGS can be applied to screen against such events. At this scale, the labour costs associated with library preparation limit the cost effectiveness of these approaches.To address these issues we seek to purchase a NextSeq500 sequencer (Illumina) alongside a BioMEK 4000 liquid handling robot that will be capable of preparing 24 independent libraries for sequencing at any one time. The NextSeq technology can theoretically sequence 96 libraries in a single run enabling cost effective high throughput sequencing of multiple bacterial genomes simultaneously. The BioMEK 4000 can also be used to prepare a wide range of libraries suitable for sequencing drastically reducing overhead costs to researchers. The cross compatibility of libraries generated between all the Illumina sequencing platforms will allow DeepSeq to prepare libraries which can be sequenced using capacity at our partner institutions.NGS technologies have a wide range of research questions. The NGS technologies outlined here will greatly enhance research at the UoN in a broad range of fields. Some of these fields include: (i) Synthetic Biology (ii) Molecular Microbiology (ii) Industrial Biotechnology and Bioenergy (iv) Food Security (v) Regenerative Medicine (vi) Stem Cell and Developmental Biology (vii) Healthy Aging (viii) Epigenetic Basis of Aging and Disease (ix) Bioscience for Health (x) Systems Biology and Bioinformatics (xi) Exploiting New Ways of Working.The development of an enhanced NGS facility at the UoN alongside the establishment of a regional Midlands Sequencing Consortium will enable the UoN and collaborators to address key biological questions in the above areas which will have a significant impact in science, the economy and society.