Our group has laid the groundwork in developing a unique, nanopore based method for DNA sequencing by nanopore induced photon emission (SNIPE), which utilizes optical detection rather than the more ubiquitous electrical detection. Our approach is superior to other nanopore approaches as the readout does not involve enzymes, parallelization is straightforward, and the readout is non-destructive. In this grant we propose three distinct aims (developed in parallel), which when brought together, will enable DNA sequencing at an unprecedented scale in terms of speed (>2 10^6 bases/s,) and extremely low cost.
Our first aim i s to dramatically increase the throughput, speed and accuracy of SNIPE. In order to achieve this, we will concentrate our efforts on parallelization of the system through arrays of nanopores (up to 100x100), transformation of the readout from 2 to 4 colors, and increasing the S/B of the readout.
Our second Aim i s to develop and optimize our proprietary DNA conversion approach, Circular DNA conversion (CDC). We plan on achieving this first though automation and optimization of CDC using a commercially available benchtop system. Post CDC optimization, we plan on developing a microfluidic device capable of converting an entire human genome.
Our third Aim i s the development of data analysis algorithms needed for base calling, consensus building, sequence assembly, and error proofing. In completing these three aims we will have achieved in developing a radically new, cost-effective DNA sequencing platform, capable of long read lengths, high speed, and high accuracy. This is expected to have a wide-ranging impact on both basic and applied biomedical research and personalized healthcare.

Public Health Relevance

The extraordinary broad impact of ultra-low cost sequencing on biomedical research, comparative genomics and cancer biology, is driving the development of a plurality of DNA sequencing methods. Our group has been developing a nanopore DNA sequencing method that utilizes optical detection from hundreds of nanopores, as the molecules are pulled electrophoretically driven through the pores. This proposal will allow us to develop this method to address the $1,000 genome challenge.

National Institute of Health (NIH)
National Human Genome Research Institute (NHGRI)
Research Project (R01)
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Special Emphasis Panel (ZHG1-HGR-N (M1))
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Schloss, Jeffery
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Boston University
Engineering (All Types)
Schools of Engineering
United States
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Squires, Allison; Atas, Evrim; Meller, Amit (2015) Nanopore sensing of individual transcription factors bound to DNA. Sci Rep 5:11643
Squires, Allison H; Atas, Evrim; Meller, Amit (2015) Genomic Pathogen Typing Using Solid-State Nanopores. PLoS One 10:e0142944
Assad, Ossama N; Di Fiori, Nicolas; Squires, Allison H et al. (2015) Two color DNA barcode detection in photoluminescence suppressed silicon nitride nanopores. Nano Lett 15:745-52
Larkin, Joseph; Henley, Robert Y; Muthukumar, Murugappan et al. (2014) High-bandwidth protein analysis using solid-state nanopores. Biophys J 106:696-704
Anderson, Brett N; Assad, Ossama N; Gilboa, Tal et al. (2014) Probing solid-state nanopores with light for the detection of unlabeled analytes. ACS Nano 8:11836-45
Squires, Allison H; Hersey, Joseph S; Grinstaff, Mark W et al. (2013) A nanopore-nanofiber mesh biosensor to control DNA translocation. J Am Chem Soc 135:16304-7
Anderson, Brett N; Muthukumar, Murugappan; Meller, Amit (2013) pH tuning of DNA translocation time through organically functionalized nanopores. ACS Nano 7:1408-14
Di Fiori, Nicolas; Squires, Allison; Bar, Daniel et al. (2013) Optoelectronic control of surface charge and translocation dynamics in solid-state nanopores. Nat Nanotechnol 8:946-51
Singer, Alon; Rapireddy, Srinivas; Ly, Danith H et al. (2012) Electronic barcoding of a viral gene at the single-molecule level. Nano Lett 12:1722-8
Atas, Evrim; Singer, Alon; Meller, Amit (2012) DNA sequencing and bar-coding using solid-state nanopores. Electrophoresis 33:3437-47

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