Application of a molecular beacon based real-time isothermal amplification (MBRTIA) technology for simultaneous detection of Bacillus cereus and Staphylococcus aureus

Abstract

A multiplex real-time isothermal amplification assay was developed using molecular beacons for the detection of Bacillus cereus and Staphylococcus aureus by targeting four important virulence genes. A correlation between targeting highly accessible DNA sequences and isothermal amplification based molecular beacon efficiency and sensitivity was demonstrated using phi(Φ)29 DNA polymerase at a constant isothermal temperature of 30°C. It was very selective and consistently detected down to 10 1 copies of DNA. The specificity and sensitivity of this assay, when tested with pure culture were high, surpassing those of currently used PCR assays for the detection of these organisms. The molecular beacon based real-time isothermal amplification (MBRTIA) assay could be carried out entirely in 96 well plates or well strips, enabling a rapid and high-throughput detection of food borne pathogens.

A modifiable microarray-based universal sensor: providing sample-to-results automation

“…Bioinformatics information gleaned from the human genome project can also provide valuable information about possible health risk factors [5]. In addition to clinical applications, nucleic acid assays have also expanded to fields such as food and beverage testing [6, 7], forensics [8, 9], and quality control in personal care products production [10]. In general, each of these applications relies upon gene amplification technology, often PCR, to first increase the total number of targets to detectable levels and then either simultaneously detect those molecular targets via “real time” fluorescence methods or at the completion of the amplification process by “end point” analysis of the resulting amplicons using DNA arrays.…”