A molecular beacon DNA microarray system for rapid detection of E. coli O157:H7 that eliminates the risk of a false negative signal

Biosens Bioelectron. 2007 Jan 15;22(6):1041-7. doi: 10.1016/j.bios.2006.04.032. Epub 2006 Jul 11.

Abstract

A DNA hybridization based optical detection platform for the detection of foodborne pathogens has been developed with virtually zero probability of the false negative signal. This portable, low-cost and real-time assaying detection platform utilizes the color changing molecular beacon as a probe for the optical detection of the target sequence. The computer-controlled detection platform exploits the target hybridization induced change of fluorescence color due to the Förster (fluorescence) resonance energy transfer (FRET) between a pair of spectrally shifted fluorophores conjugated to the opposite ends of a beacon (oligonucleotide probe). Unlike the traditional fluorophore-quencher beacon design, the presence of two fluorescence molecules allows to actively visualize both hybridized and unhybridized states of the beacon. This eliminates false negative signal detection characteristic for the fluorophore-quencher beacon where bleaching of the fluorophore or washout of a beacon is indistinguishable from the absence of the target DNA sequence. In perspective, the two-color design allows also to quantify the concentration of the target DNA in a sample down to < =1 ng/microl. The new design is suitable for simultaneous reliable detection of hundreds of DNA target sequences in one test run using a series of beacons immobilized on a single substrate in a spatial format.

Publication types

  • Evaluation Study
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • DNA, Bacterial / analysis*
  • DNA, Bacterial / genetics*
  • Equipment Design
  • Equipment Failure Analysis
  • Escherichia coli O157 / genetics*
  • Escherichia coli O157 / isolation & purification*
  • False Negative Reactions
  • Fluorescence Resonance Energy Transfer / instrumentation*
  • Fluorescence Resonance Energy Transfer / methods
  • Molecular Probe Techniques / instrumentation*
  • Oligonucleotide Array Sequence Analysis / instrumentation*
  • Oligonucleotide Array Sequence Analysis / methods
  • Reproducibility of Results
  • Sensitivity and Specificity

Substances

  • DNA, Bacterial