This proposal addresses an interdisciplinary strategy that integrates the use of highly sensitive, highly specific aptamers with nanopore-based single molecule technology. The long term goal is to develop aptamer-integrated nanopore probes for biomedical detections. Aptamers are engineered DNA/RNA that can specifically recognize broad species of proteins with high affinities. Upon binding, these powerful molecules can inhibit pathogen protein, catalyze chemical reactions, control gene expression, and regulate cellular functions. Therefore aptamers can potentially be applied as tools for exploring biological systems. Motivated by such small, but sophisticated molecules, we would like to integrate aptamer technology with our nanopore probe to construct a new generation of single molecule detector that would aid greatly in diverse disease-related detections. Nanopore technology can """"""""visually"""""""" capture the dynamic binding of a single molecule to a ligand in a nanometer-scaled pore through the discrete changes in conductance upon binding. We propose the combined use of nanopore with laboratory nanofabrication, bio-friendly surface engineering, and molecular engineering to accomplish the following specific aims: 1. Understand and manipulate various properties of nanopores for improvement of single molecule detection;2. Establish a fundamental model for single molecule detection with aptamer-equipped nanopores. Broad target species with varied aptamer types (DNA/RNA) will be tested, including Immunoglobulin E, HIV-1 reverse transcriptase, and large influenza A viral particle;3. Establishing an advanced model that employs engineered Transfer Aptamer (t-Aptamer) to mediate detection. The t-Aptamer, which composes an aptamer and a universal carrier, acts as a """"""""translator"""""""" to encode target information into the frequency of hybridization between the carrier and the immobilized receptor in the nanopore. Such a t-Aptamer-mediated detection is robust, programmable, and will ultimately lead to a universal nanopore that can discriminate different targets and perform simultaneous multi-target detection. The success of this research will greatly expand the capability of nanopores as the new generation of detection technology for biomedical analysis and high-throughput screening. Fashioning such tools is significant in nanomedicine for the quantitative characterization and precise control of molecular scale components (or nanomachinery) of living cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM079613-05
Application #
8018612
Study Section
Special Emphasis Panel (ZRG1-BCMB-H (50))
Program Officer
Edmonds, Charles G
Project Start
2007-02-01
Project End
2014-01-31
Budget Start
2011-02-01
Budget End
2014-01-31
Support Year
5
Fiscal Year
2011
Total Cost
$249,093
Indirect Cost
Name
University of Missouri-Columbia
Department
Type
Organized Research Units
DUNS #
153890272
City
Columbia
State
MO
Country
United States
Zip Code
65211
Wang, Yong; Tian, Kai; Shi, Ruicheng et al. (2017) Nanolock-Nanopore Facilitated Digital Diagnostics of Cancer Driver Mutation in Tumor Tissue. ACS Sens 2:975-981
Tian, Kai; Decker, Karl; Aksimentiev, Aleksei et al. (2017) Interference-Free Detection of Genetic Biomarkers Using Synthetic Dipole-Facilitated Nanopore Dielectrophoresis. ACS Nano 11:1204-1213
Tian, Kai; Shi, Ruicheng; Gu, Amy et al. (2017) Polycationic Probe-Guided Nanopore Single-Molecule Counter for Selective miRNA Detection. Methods Mol Biol 1632:255-268
Zhang, Xinyue; Price, Nathan E; Fang, Xi et al. (2015) Characterization of Interstrand DNA-DNA Cross-Links Using the ?-Hemolysin Protein Nanopore. ACS Nano 9:11812-9
Zhang, Xinyue; Xu, Xiaojun; Yang, Zhiyu et al. (2015) Mimicking Ribosomal Unfolding of RNA Pseudoknot in a Protein Channel. J Am Chem Soc 137:15742-52
Wang, Yong; Montana, Vedrana; GrubiĊĦi?, Vladimir et al. (2015) Nanopore sensing of botulinum toxin type B by discriminating an enzymatically cleaved Peptide from a synaptic protein synaptobrevin 2 derivative. ACS Appl Mater Interfaces 7:184-92
Zhang, Xinyue; Wang, Yong; Fricke, Brandon L et al. (2014) Programming nanopore ion flow for encoded multiplex microRNA detection. ACS Nano 8:3444-50
Wang, Yong; Tian, Kai; Hunter, Lehr L et al. (2014) Probing molecular pathways for DNA orientational trapping, unzipping and translocation in nanopores by using a tunable overhang sensor. Nanoscale 6:11372-9
Wang, Yong; Luan, Bin-Quan; Yang, Zhiyu et al. (2014) Single molecule investigation of Ag+ interactions with single cytosine-, methylcytosine- and hydroxymethylcytosine-cytosine mismatches in a nanopore. Sci Rep 4:5883
Tian, Kai; He, Zhaojian; Wang, Yong et al. (2013) Designing a polycationic probe for simultaneous enrichment and detection of microRNAs in a nanopore. ACS Nano 7:3962-9

Showing the most recent 10 out of 26 publications