Rapid identification and tracing the origin of pathogenic bacteria are imperative in response to a bioterrorist attack or an infectious disease outbreak. Pulsed-Field Gel Electrophoresis (PFGE) fingerprinting is frequently used for these tasks, particularly for tracing bacterium origins. However, the short fragment bands are lost in PFGE when extended separation times are used to resolve the long DNA molecules, thus reduce the detection specificity, because the lengths of the short fragments provide additional genotype information that could be critical to discriminate two similar genomes. Another drawback of PFGE is that the electrophoresis is too slow. In a rapid (24-hour) PFGE method the electrophoresis takes 14-18 hours. Additionally, the intra- and inter-laboratory reproducibility of PFGE also needs to be improved. Although diverse approaches such as entropic traps and DNA prisms have been explored for DNA separations, none have thus far been practically utilized. We have recently discovered a new and efficient technique to resolve broad size ranges of DNA molecules. In this application we propose to construct a novel instrument to demonstrate the proof-of-principle of this new technique for high-speed and accurate microbial identification.
(provided by applicant): We have recently discovered a new and efficient technique to resolve broad size ranges of DNA molecules. On the basis of this discovery and as a first step toward our ultimate goal of developing a system for fast microbial identification, we plan to construct a novel instrument to demonstrate the proof-of-principle of this new technique for high-speed and accurate microbial identification.
|Yu, Haiqing; Lu, Joann J; Rao, Wei et al. (2016) Capitalizing Resolving Power of Density Gradient Ultracentrifugation by Freezing and Precisely Slicing Centrifuged Solution: Enabling Identification of Complex Proteins from Mitochondria by Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass S J Anal Methods Chem 2016:8183656|
|Chen, Huang; Zhu, Zaifang; Lu, Joann Juan et al. (2015) Charging YOYO-1 on capillary wall for online DNA intercalation and integrating this approach with multiplex PCR and bare narrow capillary-hydrodynamic chromatography for online DNA analysis. Anal Chem 87:1518-22|
|Zhu, Xiaocui; Xu, Lei; Wu, Tongbo et al. (2014) Continuous monitoring of bisulfide variation in microdialysis effluents by on-line droplet-based microfluidic fluorescent sensor. Biosens Bioelectron 55:438-45|
|Wang, Wei; Gu, Congying; Lynch, Kyle B et al. (2014) High-pressure open-channel on-chip electroosmotic pump for nanoflow high performance liquid chromatography. Anal Chem 86:1958-64|
|Zhu, Zaifang; Chen, Huang; Chen, Apeng et al. (2014) Simultaneously sizing and quantitating zeptomole-level DNA at high throughput in free solution. Chemistry 20:13945-50|
|Zhu, Zaifang; Liu, Lei; Wang, Wei et al. (2013) Resolving DNA at efficiencies of more than a million plates per meter using bare narrow open capillaries without sieving matrices. Chem Commun (Camb) 49:2897-9|
|Wang, Wei; Lu, Joann J; Gu, Congying et al. (2013) Performing isoelectric focusing and simultaneous fractionation of proteins on a rotary valve followed by sodium dodecyl-polyacrylamide gel electrophoresis. Anal Chem 85:6603-7|
|Zhu, Zaifang; Chen, Huang; Wang, Wei et al. (2013) Integrated bare narrow capillary-hydrodynamic chromatographic system for free-solution DNA separation at the single-molecule level. Angew Chem Int Ed Engl 52:5612-6|