Sex selected equine embryo production system
View Patent ↗A sex selected equine embryo production system for the production of sex selected equine embryos and offspring by fertilization of oocytes with sex selected sperm by intracytoplasmic sperm injection.
1. A method of producing an equine embryo of a selected sex comprising:
(a) obtaining a plurality of equine sperm cells;
(b) determining a sex of said plurality of equine sperm cells;
(c) sorting said plurality of equine sperm cells based upon sex;
(d) obtaining an equine oocyte;
(e) intracytoplasmically injecting said equine oocyte with one of said plurality of equine sperm cells of a selected sex; and
(f) producing an equine embryo of said selected sex, wherein said equine embryo is capable of blastocyst formation.
2. The method of claim 1 , further comprising freezing said plurality of equine sperm cells sorted based upon sex.
3. The method of claim 2 , further comprising thawing said plurality of equine sperm cells.
4. The method of claim 3 , further comprising washing said plurality of equine sperm cells.
5. The method of claim 4 , wherein said washing comprises washing said plurality of equine sperm in a chemically defined medium containing heparin.
6. The method of claim 4 , further comprising swimming up said equine sperm cells prior to said washing.
7. The method of claim 6 , wherein said swimming up comprises swimming up of said equine sperm cells in condition defined medium containing about 2 mM caffeine and heparin.
8. The method of claim 1 , further comprising utilizing a piezo injection system for said injecting said one of said plurality of equine sperm cells of a selected sex into said equine oocyte.
9. The method of claim 8 , further comprising providing a sperm-injection pipette having an external diameter of about eight to ten micrometers.
10. The method of claim 1 , further comprising providing a flow cytometer which performs said determining a sex of said plurality of equine sperm cells and said sorting said plurality of equine sperm cells based upon sex.
11. The method of claim 10 , further comprising staining an amount of DNA contained within said plurality of equine sperm cells with a DNA selective material prior to said determining a sex of said plurality of equine sperm cells.
12. The method of claim 11 , further comprising exposing said plurality of equine sperm cells to a light source to generate an amount of fluorescence which varies based on the amount of said DNA staining with said DNA selective material.
13. The method of claim 12 , further comprising detecting said amount of fluorescence which varies based on the amount of said DNA stained with said DNA selective material.