Pedigree analysis of offspring.

<p>PCR amplification of genomic DNA from combs of chicks, blood from their wild-type mothers, semen from their interspecific chimeric fathers and the 169.4 PGC and 167.2 cell lines that were incorporated into the testis of the interspecific chimeric males. The GFP locus in the 169.4 and 167.2 is heterozygous and therefore, approximately one half of the offspring from the injected PGCs inherit the locus and express GFP. Top panel, offspring from the 169.4 cell line. Bottom panel; offspring from the 167.2 cell line.</p

Colonization of xenogeneic testis (Left panel) GFP positive chicken primordial germ cells in culture on irradiated BRL feeder cells.

<p>(Right panel) To evaluate the incorporation of chicken GFP positive cells into the gonads of guinea fowl, testes were retrieved, fixed in 4% paraformaldehyde and processed for cryosectioning. GFP expressing cells are present only in the seminiferous tubules suggesting that they have differentiated into spermatogonia. The absence of GFP-expressing cells in the interstitial tissue is consistent with the exclusive commitment of PGCs to the germline. Sizebar denotes 100 µm</p

Evaluation of sperm by flow cytometry and production of GFP-expressing and naked neck offspring from interspecific male guinea fowl chimeras.

<p>The GFP and naked neck loci are heterozygous and therefore, one half of the offspring from the injected PGCs inherit the expressed alleles at these loci. The high rates of fertility indicate that the testis is highly colonized by chicken PGCs in some of the interspecific male guinea fowls. Hybrids appear in the offspring of roosters with lower rates of germline transmission. GFP transmission is calculated from all embryos evaluated including embryos at D7 and chicks at hatch. Naked neck transmission was evaluated in hatched chicks and the hybrid percentage was calculated as a percentage of all offspring including both chicks and hybrids.</p

Xenogeneic offspring from guinea fowl.

<p>Upper panel: Phylogenetic relationship between chicken (Gallus domesticus) and guinea fowl (Numida meleagris). Bottom panel: primordial germ cells migrating through the blood in a 2.5 day old embryo were retrieved and cultured (A) and transfected with a GFP expression construct. GFP positive PGCs (B) were injected into guinea fowl embryos (C). The embryos were hatched (D) and males were raised to sexual maturity (E). The guinea fowl males were mated to White Leghorn females (F) and GFP positive chicken offspring (G) were obtained demonstrating that the chicken PGCs had passed through the germline of the male guinea fowl.</p