Polymerase Chain Reaction-Dynamic Light Scattering Sensor for DNA and Protein by Using Both Replication and Cleavage Properties of Taq Polymerase.

Abstract

The incorporation of AuNPs into polymerase chain reaction (PCR) has become a promising strategy to develop sensitive sensing platforms, due to desirable optical properties of AuNPs and the exponential amplification power of PCR. However, the combination of AuNPs to PCR usually fails to reach expected sensitivity along with additional steps. Here we report a one-step and universal PCR-based biosensor for size-dependent detection of nucleic acids and proteins by using the dynamic light scattering (DLS) technique. This sensing system employs a DNA modified AuNPs (AuNPs-DNA) probe to serve as the TaqMan like signal probe, in which DNA on the surface of AuNPs can be cleaved by Taq polymerase during the extension process of PCR, causing the aggregation of AuNPs to be detected by DLS. This strategy enables sequence-specific recognition of target double-stranded DNA (dsDNA), overcoming the background signal change that triggered by the increase of the PCR cycle. It further demonstrates its applicability in detecting a foodborne pathogen Listeria monocytogenes with a detection limit of 1.2 fg μL-1, which is more sensitive than colorimetric methods. Moreover, by utilizing a proximity assay strategy, this biosensor shows the capability to the homogeneous detection of protein, showing a detection limit of 1.0 pM for a model thrombin. Together, the work demonstrates a reliable strategy to incorporate AuNPs into PCR amplification process as a signal probe, instead of the post-PCR process.