We describe a revolutionary methodology for rapid and massively parallel DNA sequencing that promises to considerably reduce the time and cost of genome sequencing. Our method includes two main steps: 1. Conversion of the target DNA molecules into easily readable code units (Design Polymers); and 2. rapid readout of the design polymers using a nanopore based approach. The first step, which has been recently demonstrated by Lingvitae AS, is described in more detail in a separately submitted related R21 proposal. Here we present a novel readout platform based on the simultaneous optical probing of multiple nanopores. ? ? The unique combination of Designed Polymers with the nanopore optical readout platform, eliminates the uncertainties associated with the development of new chemical compounds, required in other approaches. Moreover, because the nanopore readout does not rely on the enzymatic incorporation of nucleotides, much higher DNA readout speeds are expected, making our method cheaper and faster. In this proposal we lay down a straightforward experimental strategy for testing our approach. This strategy relies heavily on our expertise in nanopores and in optical probing of single-biomolecules. ? ? The specific aims of our proposal are: A. fabrication of an instrument for concurrent electrical and optical probing of DNA molecules inside the nanopore (nearly completed); B. testing the DNA readout of 5, 10 and 20 nucleotide single-stranded DNA using our nanopore setup; and, C. testing DNA readout from multiple pores simultaneously. We expect that the program that we propose will enable us to complete the development of the method within a period of two years. ? ?
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