Alternative reproductive tactics are discrete, intrasexual differences in reproductive behavior within a population. In some cases,  these complex phenotypes are determined by autosomal supergenes or sex chromosomes—both of which exhibit reduced recombination and thus enable the linked inheritance of co-adapted alleles from multiple loci. Most alternative reproductive tactics in amphibians are plastic (and reversible), environmentally determined, and lacking morphological differentiation, but a striking exception is found in the two-lined salamander (Eurycea bislineata) species complex. In some populations, two distinct male phenotypes coexist: “searching” males have mental glands, protruding premaxillary teeth, and elongate cirri used in terrestrial courtship, while “guarding” males lack these traits and instead have hypertrophied jaw musculature used in mate-guarding at aquatic nesting sites. These tactics differ in many morphological and behavioral phenotypes, but their proximate cause has not yet been described. Here, we generated genome-wide SNP data from >130 Eurycea cf. wilderae collected from Highlands Biological Station. We provide evidence for a XY sex determination and for a Y-linked polymorphism underlying alternative reproductive tactics in this population. We then develop and validate a PCR-based genotyping assay and apply this to characterize the sex ratio and relative frequency of tactics from a sample of larvae. Our results add to the growing body of literature exploring the importance of supergenes and sex chromosomes in complex intraspecific polymorphisms, and we highlight opportunities for future work to continue exploring the genomic architecture of these traits.