Sexing of Embryos:
Sexing of embryos before transfer is especially sought by the dairy cattle industry where females are the desired milk producing unit. To be useful, sexing techniques must be accurate, efficient, rapid and without detrimental effects on the embryos. Embryos can be sexed by use of antibodies to male specific antigens such as the H-Y antigen. When a fluorescent second antibody approach is used, this method is non-damaging to embryos and approximately 85% accurate for cattle embryos (White et al., 1987; Wachtel et al., 1988; White, 1988).
Recently highly accurate methods for sexing embryos which use only a few cells and provide a quick answer have been developed. These methods involve the use of Y chromosome-specific bands on a gel from a labeled homologous recognition probe.
Y-specific fragments are used as probes to locate homologous sequences present in DNA from blastomeres or trophoblast cells of embryos. As few as two to five blastomeres can be biopsied from embryos and, using an oligonucleotide polymerase chain reaction for signal amplification, embryonic sex can be determined in 6 hrs or less. Several Y specific probes are currently available for sex selection of cattle embryos (Leonard et al., 1987; Ellis et al., 1988; Popescu et al., 1988; Reed et al., 1988; Bondioli et al., 1989; Herr et al., 1989a,b, 1990) and one has been developed for swine. Recently the Y chromosome deletion assay has been simplified to a cow side field assay kit of excellent accuracy (Herr et al., 1990b,c).
While the sexing of embryos provides a way to predetermine sex, the commercial production of offspring of a chosen sex would be greatly facilitated if sperm could be sexed and the sexed semen distributed through artificial insemination. A dairyman may wish all females while specific superior cows might be inseminated with male sperm to produce beef or dairy bulls. Until recently the prospects for sexing semen have been dim. It has been known for some time that sperm containing either an X or a Y chromosome could be separated by fluorescence of stained DNA of X and Y sperm using a fluorescence activated cell sorter. This became a standard confirmatory assay but the sperm were dead after the separation. Recently Johnson et al., (1989) have reported a modification of this method wherein sperm remain alive. When female rabbits were inseminated with sorted sperm from the X chromosome population 94% of the offspring born were females. After insemination of the Y bearing sperm 81% of the offspring were male. Because of the large sample required for insemination there was overlap between the base of the X and Y peaks. If in vitro fertilization and small sperm numbers were used it should be possible to use only the completely separated peaks of X and Y sorted populations thereby increasing the accuracy to nearly 100%. Commercial use through artificial insemination will be limited until more efficient sorting systems are developed and damaging effects of the fluorescence staining on chromatin will need to be prevented.