The authors describe the use of gold-decorated magnetic nanoparticles (Au/MNPs) in discriminating DNA sequences with a single-base (guanine) mismatch. The Au/MNPs were characterized through dynamic light scattering, X-ray diffraction, superconducting quantum interference device, and UV/visible spectroscopy. They were then conjugated to a probe oligomer consisting of a hairpin-shaped DNA sequence carrying two signalling fluorophores: fluorescein at its 3' end and pyrene in the loop region. When interacting with the target DNA sequences, the hybridized probe-target duplex renders the pyrene signal (at excitation/emission wavelengths of 345/375 nm) either quenched or unquenched. Quenching (or nonquenching) of the pyrene fluorescence depends on the presence of a guanine (or a nonguanine) nucleotide at the designated polymorphic site. The linear range of hybridization in these Au/MNPs is from 0.1 nM to 1.0 μM of ssDNA. Conceivably, this system may serve as a single-nucleotide polymorphism probe. Graphical Abstract Schematic presentation of probe-conjugated Au/MNP preparation (upper panel) and working principle to discriminate DNA with or without single-base (guanine) mismatch sequences at the polymorphic sites (lower panel). Py denotes pyrene-hooked pyrrolocytidine; F denotes fluorescein.