A C-nucleoside analog having pyren-1-ylpyridine as the base moiety has been synthesized and incorporated in the middle of a short oligodeoxynucleotide. A portion of this oligonucleotide is cyclopalladated at the modified residue, and the potential of both the metal-free and the palladacyclic oligonucleotide as hybridization probes for single-nucleotide polymorphism genotyping is assessed by melting studies on relevant duplexes using various techniques. Conventional ultraviolet (UV) melting profiles at 260 nm reveal considerable destabilization of the palladacyclic duplexes relative to their metal-free counterparts. Circular dichroism melting temperatures are higher than their UV counterparts, especially with the palladacyclic duplexes. Cyclopalladation markedly reduces the fluorescence emission of the pyrenylpyridine moiety, but both the metal-free and the palladacyclic oligonucleotide exhibit a qualitatively similar pattern of increased fluorescence on hybridization with a complementary sequence, consistent with the pyrene ring being “pushed out” of the base stack. Emission at low temperature is dependent on the nucleobase paired with the pyrenylpyridine base surrogate with both of the modified oligonucleotides. This discrimination is stronger with the palladacyclic oligonucleotide, possibly owing to Pd(II)-mediated base pairing.