Animal husbandry – Avian egg treatment or production
Reexamination Certificate
2000-03-23
2002-03-12
Jordan, Charles T. (Department: 3644)
Animal husbandry
Avian egg treatment or production
C424S582000
Reexamination Certificate
active
06354242
ABSTRACT:
FIELD OF THE INVENTION
The present invention concerns methods of transferring primordial germ cells to birds for the production of gametes therein. Such methods are useful in the conservation of endangered avian species, in reducing the time required to produce spermatozoa from slowly maturing species such as turkeys, decreasing the costs of maintaining breeder flocks, and altering the sex ratio of offspring flocks (e.g., to enhance the efficiency of production).
BACKGROUND OF THE INVENTION
The ability to more easily produce gametes of particular avian species would be extremely useful to the avian veterinary and poultry production fields. For endangered species such as the whooping crane, it would be extremely useful to have a ready supply of male spermatozoa. For commercial birds such as turkeys, it would be desirable to more quickly and economically produce male spermatozoa. For meat- producing flocks, it is desirable to have ways to increase the ratio of male birds in the flock. Accordingly, there is a need for new ways to obtain avian spermatozoa.
Chimeras are composite organisms consisting of cells derived from more than one zygote. Experimental chimeras have been used to study cell to cell interaction and cell lineage analysis during development (A. McLaren,
Mammalian Chimeras
. Cambridge University Press, Cambridge (1976)). When chimeras are produced using material derived from very early embryos, organisms develop containing a full mixture of somatic tissues. If the starting material includes early germ cells or their precursors, the resulting individuals will produce gametes of both the donor and recipient genotypes. In addition, chimeras can be intraspecific, i.e. between two zygotes of the same species, or interspecific, i.e. between two different species.
Avian primordial germ cells (PGCs) like other vertebrate germ cells are extragonadal in origin and must undergo a complex journey to reach the gonad. The transfer of blastodermal cells and primordial germ cells has produced avian germline chimeras.
Reynaud (
J. Embryol. Exp. Morphol
. 21:485-507 (1969)), a pioneer in the production of avian germline chimeras, reported the production of turkey-chicken germline chimeras by the intravascular transfer of dissociated turkey germinal crescent cells into previously sterilized chick embryos (accomplished by exposure of the recipient germinal crescent to ultra-violet light). PGCs obtained by mechanical dissociation of the endoderm of the germinal crescent (stage 5) were injected into the blood vessels of chicken embryos (3-5 days of incubation). Prior to injection the recipient embryos were sterilized at stage 8-10 (H&H) with ultraviolet light; however, the sterilization was not complete. The turkey PGCs in the chick embryo were identified solely on the basis of their nucleoplasmic ratio. This method of identification was difficult and tenuous and could not be used for actively dividing turkey PGCs since the dividing germ cells gave an aberrant nucleoplasmic ratio. In a succeeding study, the transferred PGCs were allowed to undergo maturation in the host gonads and apparently could give rise to gametes but they were not suitable for fertilization (Wilhelm Roux
Arch. Dev. Bio
. 179:85-110 (1976)). The spermatozoa were incapable of fertilizing turkey eggs. They fertilized chick eggs but there was no normal development. Chicken spermatozoa were capable of activating the eggs obtained from female interspecific chimeras but they did not give rise to embryos. When the eggs were fertilized by turkey spermatozoa they developed into abnormal embryos that did not survive beyond stage 38 (H&H). Reynaud used morphology as the only distinguishing characteristic in an attempt to identify turkey germ cells from chicken germ cells. Morphology alone is not sufficient for identifying chimeras and must be substantiated with other markers. In addition, according to Aige-Gil and Simkiss (
Brit. Poul. Sci
. 32:427-438 (1991)), the presence of turkey gametes was not identified by test matings. Accordingly, there remains a need for new ways to accomplish the production and transfer of avian gametes.
SUMMARY OF THE INVENTION
A method for the production and collection of avian sperm comprises the steps of: providing primordial germ cells from a donor avian species; administering the primordial germ cells to a recipient avian species in ovo; incubating the recipient avian species to hatch; and then collecting sperm of the donor avian species from the recipient avian species. For example, the donor avian species may be a whooping crane, and the recipient avian species may be a sand hill crane. In another example, the donor avian species may be a turkey, and the recipient avian species may be a chicken.
In birds, unlike mammals, it is the male that is the homogametic sex (ZZ) and the female which is the heterogametic sex (Zw). Therefore in birds, it is the female that determines the gender of the offspring since she produces ova which carry either the Z or w chromosome. Thus, as noted below, by transferring male primordial germ cells to female embryonic hosts, the percentage of Z-bearing ova produced by that host is increased and the percentage of male offspring is increased. An increase in the percentage of male offspring from broiler flocks is economically desirable for the corresponding greater feed conversion ratio and more efficient meat production so obtained.
Accordingly, a second aspect of the present invention is a method of increasing the number of male birds hatched from a plurality of bird eggs, comprising the steps of: administering to a female bird in ovo male (ZZ) avian primordial germ cells; incubating the female bird to hatch; raising the female bird to sexual maturity; and then breeding the bird to produce a plurality of fertile bird eggs (with the ratio of male to female birds eggs produced from the bird being greater than that obtained in the absence of administering the male primordial germ cells to the bird in ovo.) Typically, the method further comprises the step of incubating the plurality of bird eggs to hatch (with the ratio of male to female birds produced from the plurality of eggs being greater than that produced in the absence of administering the male primordial germ cells to the female bird in ovo). The female bird may be of any suitable species, such as chicken or turkey, and the primordial germ cells being administered are preferably from the same species as the female bird to which they are administered.
The foregoing and other objects and aspects of the present invention are explained in detail in the specification set forth below.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
“Bird” or “avian species” as used herein refers to any avian species, including but not limited to chicken, turkey, duck, geese, quail, pheasant, and ostrich. Any of numerous other species can be employed to carry out the present invention, particularly when it is used for the conservation of endangered species such as the whooping crane (where the recipient species would be the sand hill crane). “Egg” as used herein refers to avian eggs that contain live embryonic birds. “Primordial germ cell” or “PGC” as used herein refers to the most differentiated diploid cell line in the embryo that will ultimately develop into haploid gametes (either sperm or ova).
“SSEA-1 antibody” refers to an antibody, preferably a monoclonal antibody, that specifically binds to the stage specific embryonic antigen-1 (SSEA-1) (M. Buehr
Exp. Cell Res
. 232, 194-207 (1997)). SSEA-1 is a carbohydrate epitope determined by galactose &bgr;1→4 fucose &agr;1→3 N acetylglucosamine linkage (H. Gooi et al.,
Nature
292, 156-158 (1981)). A monoclonal antibody to SSEA-1 was developed by the fusion of mouse myeloma cells with spleen cells from a mouse that had been immunized with F9 teratocarcinoma cells (D. Solter and B. Knowles,
Proc. Natl. Acad. Sci. USA
75, 5565-5569 (1978)). SSEA-1 antibody is known as an avian immunohistochemical germ cell marker (L. Karagenc et al.,
Dev. Genet
. 19, 290-301 (1996)). P
D'Costa Susan
Pardue Samuel L.
Petitte James N.
Abbott Yvonne R.
Jordan Charles T.
Myers Bigel & Sibley & Sajovec
North Carolina State University
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