Nonenzymatic Detection of Bacterial Genomic DNA Using the Bio Bar Code Assay

Abstract

The detection of bacterial genomic DNA through a nonenzymatic nanomaterials-based amplification method, the bio bar code assay, is reported. The assay utilizes oligonucleotide-functionalized magnetic microparticles to capture the target of interest from the sample. A critical step in the new assay involves the use of blocking oligonucleotides during heat denaturation of the double-stranded DNA. These blockers bind to specific regions of the target DNA upon cooling and prevent the duplex DNA from rehybridizing, which allows the particle probes to bind. Following target isolation using the magnetic particles, oligonucleotide-functionalized gold nanoparticles act as target recognition agents. The oligonucleotides on the nanoparticle (bar codes) act as amplification surrogates. The bar codes are then detected using the Scanometric method. The limit of detection for this assay was determined to be 2.5 fM, and this is the first demonstration of a bar code-type assay for the detection of double-stranded, genomic DNA.