Germline chimeric chickens from dispersed donor blastodermal cells and compromised recipient embryos

ABSTRACT Stage-X blastoderms, within intact eggs from White Leghorn hens, were exposed to 500-700 rads of radiation from a 60Co source prior to injection, into the subgerminal cavity, of approximately 100 or 200-400 dispersed cells from stage-X blastoderms isolated from eggs laid by Barred Plymouth Rock hens. Embryos developing past day 14 of incubation and hatched chicks were assessed for donor and recipient cell contribution to the melanocyte population through examination of black and yellow down pigmentation, respectively (Barred Plymouth Rocks have a recessive allele at the I locus while the White Leghorns have a dominant allele at the I locus). Of the 809 embryos injected with approximately 100 cells, 192 developed past day 14 and black pigmentation, indicating somatic chimerism, was observed on 118 of the 192 (58%) embryos and chicks. Of the 296 embryos injected with 200-400 donor cells, 86 developed past day 14 of incubation. Somatic chimerism was observed on 55 of the 86 (64%) embryos and chicks. To test for germline chimerism, birds surviving to maturity were mated to Barred Plymouth Rocks. Five somatically chimeric females were produced when approximately 100 cells were injected, and one was a germline chimera. Six somatic female chimeras were produced following the injection of 200-400 cells, three of which proved to be germline chimeras by the presence of Barred Rock chicks among their offspring. Two of the nine males produced by injecting approximately 100 cells were germline chimeras. Five of 6 somatic male chimeras, produced by injection of 200-400 cells, have sired black chicks; furthermore, one of the roosters produced by the injection of 200-400 cells has sired only donor-derived chicks. These data indicate that irradiation of the recipient embryo, prior to injection of the donor cells, consistently yields somatic and germline chimeric chickens. The ability to insert donor cells into the germline provides a powerful new tool that will facilitate molecular and cellular manipulation of the developing chick embryo.