Preserving poultry and avian biodiversity needs the development of new techniques cryoconserve bird species. Avian gene preservation research has recently focused on the use of the early precursors of the reproductive cells, the primordial germ cells. This is because avian PGCs have a unique migration route through the vascular system which offers easy accessibility in the young avian embryo. The few germ isolated germ cells can be cultured in vitro, frozen, reintegration into a surrogate host recipient embryo and raised to sexual maturity. The efficient recovery of the donor genotype and the frequency of germline transmission from the surrogate host animals are still areas which need further investigation. Here, we will tell you our efforts and successes in the cryopreservation and reestablishment of local chicken breeds in both France and Hungary. Recently we have developed in France reproductive biotechnologies based on PGCs using as a model a local breed La Poule Noire du Berry (NB), which have benefited from a national program of conservation. The in vitro steps that PGCs undergo before their use for reproduction may affect their integrity and reproductive capacities. In this study we investigated the effect of the culture duration and cryopreservation (comprising cell freezing, thaw and reamplification in vitro) on the gene expression, DNA methylation, and germline transmission rate of NB PGCs. RNAseq study and RRBs analysis revealed strong gender differential effect of long term in vitro cultures on transcript abundances and DNA methylation of PGCs. Cryopreservation affected DNA methylation in PGCs of both sexes, but not their gene expression. We obtained for the first time in France the progeny from derived in vitro and cryopreserved PGCs. The germline transmission rate for female and male NB PGCs was up to 60.6% and up to 42.4% respectively, and seemed to vary according to the culture duration in a gender-specific manner. These results will serve to better master in vitro culture conditions and improve germline transmission rate of avian PGCs.