Long-term rock dove (Columba livia) primordial germ cell culture: A tool towards avian conservation.

Primordial germ cells (PGCs) are critical tools for genome engineering and conservation in birds. Although culture systems for chicken PGCs have been well established for nearly two decades, efforts to propagate PGCs from other avian species have proved exceptionally challenging, limiting the broader application of artificial reproductive technologies in birds. Here we report the first successful derivation and long-term culture of PGCs from the rock dove, or common pigeon (Columba livia). Guided by transcriptomic profiling of PGCs, we developed a species-specific medium that supports PGC maintenance and expansion. We identify insulin signaling as a requirement for survival under the conditions tested, and demonstrate that inhibition of the retinoic acid receptor, in the presence of vitamin A, supports propagation. Supplementation with bone morphogenetic protein 4, leukemia inhibitory factor (LIF), glial cell line-derived neurotrophic factor, and pleiotrophin further enhances PGC proliferation. Cultured cells express canonical germline markers and migrate to the gonads following injection into both rock dove and chicken embryos, confirming functional migratory competency. These findings establish a platform for germline manipulation and biobanking in Columbidae, broadening the potential applicability of reproductive technologies to conservation efforts.