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Enzyme-less DNA base discrimination using solid-state nanopores with high-frequency integrated detection electronics

Award Number: R01HG009189
ORGANIZATION: NATIONAL HUMAN GENOME RESEARCH INSTITUTE
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

Group Awards By Issue Date FY or Funding FY:

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Enzyme-less DNA base discrimination using solid-state nanopores with high-frequency integrated de- tection electronics There is strong demand for third-generation DNA sequencing systems to be single-molecule, massively- parallel, and real-time, while also reducing operating costs and supporting long read lengths. No technologies have yet met this challenge, but the most successful attempts to date have been based on methods which track the real-time operation of single enzyme molecules operating on a strand of DNA. Optical approaches to single-polymerase imaging suffer from low signal-to-noise ratios deriving from the weak photon emission from single fluorophores (< 2500 photons/sec), and thus demand both complex optics and purposely-reduced base incorporation rates (~1 Hz). Nanopore-based detection approaches offer faster detection and have been demonstrated to track polymerase activity at higher incorporation rates (~10-100 Hz), but have struggled with reliability issues and high error rates associated with the still-weak signal levels produced by popular protein nanopores. These struggles suggest that nanopore-based single-molecule sequencing techniques which do not de- pend on real-time imaging of active enzymes would have several important advantages. First and foremost, they could offer sequencing speeds even faster than a free-running polymerase molecule (which operate at ~100-1000 Hz). Second, removing active enzymes from the detection platform offers more freedom to optimize key parameters such as buffer conditions and temperatures outside the operating range of natural enzymes. Third, sequencing platforms without active enzymes may prove simpler and cheaper to operate, ship, and store. Lastly, nanopores, particularly biological ones, face reliability challenges as electronic devices, experi- encing degradation during use. In this three-year effort, we focus on the development of a multiplexed solid-state nanopore platform ena- bling a per-pore sequencing rate of at least 105 bases/sec at an error rate of less than 0.1%, leveraging inte- grated electronics and state-of-the-art solid-state nanopores based on ultra-thin membrance and delivering useful signal bandwidths in excess of 10 MHz when required. We expect to be able to delect signal levels as low as 225 pA at signal-to-noise ratios greater than 8 and bandwidth better than 3 MHz, making possible high- speed free-running single-molecule electrophoretic sequencing. This goal is pursued through three Specific Aims: the design of biomimetic solid-state nanopores with sensing geometries driven to single nanometer siz- es (Specific Aim 1), the design of electronics optimized for high-speed multiplexed detection of these na- nopores (Specific Aim 2), and data analysis of the resulting signals (Specific Aim 3).


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2023 ( Subtotal = $0 )
2023	2018	THE TRUSTEES OF COLUMBIA UNIVERSITY IN THE CITY OF NEW YORK	615 W 131ST ST	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	3	11/18/2022	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2021 ( Subtotal = $0 )
2021	2018	TRUSTEES OF COLUMBIA UNIVERSITY IN THE CITY OF NEW YORK, THE	116TH AND BDWY	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	3	3/29/2021	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2020 ( Subtotal = $0 )
2020	2018	TRUSTEES OF COLUMBIA UNIVERSITY IN THE CITY OF NEW YORK, THE	116TH AND BDWY	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	3	5/4/2020	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2018 ( Subtotal = $352,905 )
2018	2018	TRUSTEES OF COLUMBIA UNIVERSITY IN THE CITY OF NEW YORK, THE	116TH AND BDWY	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	3	7/16/2018	NON-COMPETING CONTINUATION	$352,905
														Subtotal = $352,905

Issue Date FY: 2017 ( Subtotal = $358,580 )
2017	2017	TRUSTEES OF COLUMBIA UNIVERSITY IN THE CITY OF NEW YORK, THE	116TH AND BDWY	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	2	7/6/2017	NON-COMPETING CONTINUATION	$358,580
														Subtotal = $358,580

Issue Date FY: 2016 ( Subtotal = $461,366 )
2016	2016	COLUMBIA UNIVERSITY	615 WEST 131ST ST, 4TH FLOOR	NEW YORK	NY	10027	NEW YORK	USA	Human Genome Research	000	1	9/16/2016	NEW	$461,366
														Subtotal = $461,366

Grand Total All Awards = $1,172,851

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Enzyme-less DNA base discrimination using solid-state nanopores with high-frequency integrated detection electronics

Award Number: R01HG009189

ORGANIZATION: NATIONAL HUMAN GENOME RESEARCH INSTITUTE

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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