Exonuclease Epigenetic Sequencing

Project Summary There is a current need within the field of next generation sequencing (NGS) for new, enabling instrumentation, capable of high accuracy, direct, native DNA sequencing, including the identification of canonical and modified bases and the correct characterization of homopolymer stretches and repeating sequences. During this program, Electronic BioSciences (EBS) and a team of field experts aim to solve the technical challenges associated with the development of a completely new and novel nanopore-based sequencing platform, including the associated methodology for sequencing DNA at the single nucleotide level, with the capability of directly and correctly identifying chemically modified nucleotides. During this project, efforts will specifically focus on the high accuracy detection/identification of 5-methylcytosine (5mC) and N6-methyladenine (m6A) sequencing, before pursuing other modifications. At present, the scientific community’s understanding of the “epigenome,” i.e. the chemical modifications which regulate the function of DNA, is still in its infancy. While there are many known chemical modifications to either the base or sugar-phosphate backbone of nucleic acids, due to the lack of analytical characterization methods available, the exact roles of these modifications remain to be assessed. New technologies capable of elucidating the roles of these modifications have the potential to revolutionize the use of the epigenome. Furthermore, with regards to homopolymer and repeat sequences of andgt;6 nucleotides which are commonly found in genomes, clinicians have identified that many of these sequences are expanded, contracted, or mutated in cancers, neurological disorders, and heritable diseases, and therefore, sequencing for changes in these regions has promising potential for utilization as routine genetic markers for diagnostics and prognostics purposes. At the conclusion of this Fast-Track project, a multiplexed sequencing instrument that is ready for immediate, expanded, initial user base use in laboratory settings will be developed and built, and complete concept feasibility will have been demonstrated through both synthetic and genomic DNA sequencing, including 5mC and m6A characterization.Project Narrative The direct sequencing of chemical modifications in the genome with high accuracy would facilitate an explosion of epigenetic sequencing data that has otherwise not been attainable to date. Thus, the methodology/technology that will be developed during this project has the potential to revolutionize the use and understanding of the genome and epigenome. More long-term, these results would better enable the use of epigenetic markers in clinical diagnostics and prognostics, provide a deeper understanding of infectious diseases, allow cultivation of better crops, and advance our basic understanding of all domains of biology, in addition to empowering unforeseen fields, such as epigenomic diagnostics and therapeutics.