Primordial germ cells (PGCs) are the precursors of sperm and eggs1. In most animals, segregation of the germ line from the somatic lineages is one of the earliest events in development2; in avian embryos, PGCs are first identified in an extra-embryonic region, the germinal crescent, after approximately 18 h of incubation. After 50–55 h of development, PGCs migrate to the gonad and subsequently produce functional sperm and oocytes3,4. So far, cultures of PGCs that remain restricted to the germ line have not been reported in any species5,6. Here we show that chicken PGCs can be isolated, cultured and genetically modified while maintaining their commitment to the germ line. Furthermore, we show that chicken PGCs can be induced in vitro to differentiate into embryonic germ cells that contribute to somatic tissues. Retention of the commitment of PGCs to the germ line after extended periods in culture and after genetic modification combined with their capacity to acquire somatic competence in vitro provides a new model for developmental biology. The utility of the model is enhanced by the accessibility of the avian embryo, which facilitates access to the earliest stages of development and supplies a facile route for the reintroduction of PGCs into the embryonic vasculature. In addition, these attributes create new opportunities to manipulate the genome of chickens for agricultural and pharmaceutical applications.
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We thank C. Gitter for technical assistance with the karyotype; A. Pradas-Monne for help in the laboratory; W. Halfter for providing the 1B3 antibody; Leica for the provision of optical equipment to photograph the GFP-positive embryo; and J.-M. Buerstedde for supplying β-actin-neo and β-actin-puro. This work was supported by the Small Business Innovation Research Programs of the USDA and the NIH to Origen Therapeutics and a USDA grant to M.E.D. Author Contributions M.C.L. developed the cell culture system with the assistance of J.H.D., P.A.L. and R.B.; C.M.-L. and J.H.D. performed the embryological manipulations; P.A.L. executed the molecular biology in collaboration with B.S.H. and L.T.H.; A.K. provided animal care; T.M.G., S.E.S. and M.E.D. conducted the telomerase assay and karyotyping; M.C.L. and R.J.E. coordinated the contributions of authors and wrote the paper. All authors discussed the results and commented on the manuscript.
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Efficient production of human interferon beta in the white of eggs from ovalbumin gene–targeted hens
Scientific Reports (2018)