710191Population Genetics: Absolute and Relative Quantification of DNA Using Molecular CountingClosedGRD Proof of ConceptInnovate UKAlthough many different techniques are available to measure amounts of DNA, each has tradeoffs with respect to sensitivity, selectivity, dynamic range, convenience, robustness and cost. Digital methods (digital PCR [dPCR], NanoString, single-molecule sequencing) are gradually superseding analog alternatives (quantitative PCR [qPCR], microarrays) because of inherent statistical advantages. Population Genetics (PG), and others, have recently described new digital counting methods based on next-generation sequencing (NGS) termed ‘CounterStamping’. DNA molecules in a sample of interest are labelled with a DNA counter sequence from a large repertoire of counter sequences. Sequencing of the counter and its attached labelled molecule can provide information on the relative abundance of different molecular species and estimate the absolute molecule number. The generated data thereby allow quantification of both known and previously unidentified genetic variation, unlike existing methods that can generally only quantify known variation. Despite significant benefits, use of counting methods is hampered because existing protocols are complicated and inconvenient compared to qPCR/dPCR. This proposal is to develop proof of concept to streamline and simplify PCR-based counting for absolute and relative quantification. PG’s proposed method is quicker, easier and has reduced possibility of contamination compared to current approaches. PG also proposes to develop algorithms to correct errors in counter sequences to improve the accuracy of the method. Continuing reductions in NGS costs, and increased adoption of desktop sequencers, make counting PCR potentially disruptive to existing qPCR and dPCR technologies. Further, the proposed technology will drive new applications in disease monitoring, companion diagnostics and molecular diagnostics where greater multiplexing and precise measurements are required.