Jingkang Huang

Primordial germ cells (PGCs) are the earliest precursors of gametes and essential cellular materials for poultry germplasm conservation and genetic modification. In this study, PGCs isolated from 2.5-day male and female Silkie chicken embryos were defined as circulating PGCs (cPGC_M and cPGC_F), whereas PGCs isolated from 8.5-day male and female embryos were defined as gonad-derived PGCs (gPGC_M and gPGC_F). RNA sequencing with three biological replicates per group was performed to characterize developmental-stage- and sex-associated transcriptional programs. Comparative transcriptomic analyses identified stage-associated differences in pathways related to cell–matrix interaction, focal adhesion, PI3K–Akt signaling, and stem cell pluripotency, with more differentially expressed genes detected in the female than in the male stage comparison. Sex-biased expression was detectable at the circulating stage, mainly reflecting W- and Z-linked expression differences, and the number of sex-biased genes was greater after gonadal colonization, primarily because more autosomal differentially expressed genes were detected. Candidate genes and pathways, including HINTW, DMRT1, SOX2, EDNRB, LPAR4, GFRA3, ECM–receptor interaction, ribosome-related modules, and calcium signaling, were associated with these comparisons and are presented as targets for future validation rather than as confirmed regulators. Because the study used three biological replicates per group, RT-qPCR corroboration was limited to six female stage-associated genes, and no functional perturbation assays were performed, the findings are descriptive and do not establish causal mechanisms of PGC migration, gonadal colonization, or sex differentiation. Within these limitations, the dataset provides an exploratory resource for subsequent studies of chicken PGC biology and biotechnology.