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Methods for Single-Molecule Sensing and Detection Using Bacteriophage Phi29 DNA Packaging Motor. | LitMetric

Methods for Single-Molecule Sensing and Detection Using Bacteriophage Phi29 DNA Packaging Motor.

Methods Mol Biol

Division of Pharmaceutics and Pharmaceutical Chemistry, College of Pharmacy, The Ohio State University, Columbus, OH, USA.

Published: March 2019

AI Article Synopsis

  • * Advanced biophysical methods like single molecule photobleaching and fluorescence resonance energy transfer are used to study the motor components, allowing researchers to analyze their structure and function at an individual molecule level.
  • * The phi29 system's ability to be engineered opens up exciting possibilities for applications in fields like nanotechnology and nanomedicine, including DNA sequencing, targeted drug delivery, and disease diagnostics.

Article Abstract

Bacteriophage phi29 DNA packaging motor consists of a dodecameric portal channel protein complex termed connector that allows transportation of genomic dsDNA and a hexameric packaging RNA (pRNA) ring to gear the motor. The elegant design of the portal protein has facilitated its applications for real-time single-molecule detection of biopolymers and chemicals with high sensitivity and selectivity. The robust self-assembly property of the pRNA has enabled biophysical studies of the motor complex to determine the stoichiometry and structure/folding of the pRNA at single-molecule level. This chapter focuses on biophysical and analytical methods for studying the phi29 motor components at the single-molecule level, such as single channel conductance assays of membrane-embedded connectors; single molecule photobleaching (SMPB) assay for determining the stoichiometry of phi29 motor components; fluorescence resonance energy transfer (FRET) assay for determining the structure and folding of pRNA; atomic force microscopy (AFM) for imaging pRNA nanoparticles of various size, shape, and stoichiometry; and bright-field microscopy with magnetomechanical system for direct visualization of viral DNA packaging process. The phi29 system with explicit engineering capability has incredible potentials for diverse applications in nanotechnology and nanomedicine including, but not limited to, DNA sequencing, drug delivery to diseased cells, environmental surveillance, and early disease diagnosis.

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-4939-8556-2_21DOI Listing

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