Sex determination of Syrian hamster pups using placental tissues

Abstract

Molecular sex determination in Syrian hamsters (Mesocricetus auratus) has been limited by incomplete annotation of Y-linked loci in currently available genome assemblies. Here, we evaluate the Y-linked gene PRSSLY, which encodes a testis-specific serine protease-like protein, as a molecular marker for genetic sexing of Syrian hamster embryonic and placental tissues. Primers flanking a conserved PRSSLY coding region produced a male-specific amplicon showing 100% concordance with results from the established KDM5C/KDM5D PCR assay in E15.5 tail biopsies. SYBR Green-based qPCR enables the accurate detection of PRSSLY, characterized by a unique melt-curve profile, exclusively in male samples, allowing for efficient and sensitive mid-throughput analysis. Among 417 placental samples from 39 dams tested, which naturally contain a mixture of maternal and embryonic genomes, the sex of individual pups was classified with 85% specificity, with the remaining ambiguous samples confirmed as male by PCR and agarose gel electrophoresis. This assay provides a robust and reproducible approach for accurate sex genotyping in developmental and reproductive studies using Syrian hamsters.