A PCR-free, optics-free platform has been developed for the detection of bacterial 16S rRNA at 1 aM (10 M). This sequence-specific nucleic acid (NA) detector is based on electromechanical signal transduction using a nanopore-based scheme. Our device utilizes peptide nucleic acid (PNA) capture probes conjugated to polystyrene beads. Since PNA is charge neutral, the bead-PNA conjugates may be designed to be charge neutral until they hybridize target NA, at which point the complex becomes negatively charged and mobile in an electric field. If the electric field is oriented through a pore that is too small for the bead-PNA conjugate with hybridized target to pass through, it will block the pore thereby causing an easily measured step reduction in ionic current. In this way, the selective, NA hybridization event is electromechanically transduced. This device has proven capable of detecting E. coli 16S rRNA at 1 aM against a 1 pM background of RNA from Pseudomonas putida and shows promise for integration into portable, low-cost systems for rapid detection of pathogenic bacteria in body fluids, food, and water.
Sequence-Specific Nucleic Acid Detection at 1 aM
A presentation by Harold Monbouquette