The objectives of the proposed research are part of an overall program to identify the critical molecular phenomena underlying early mammalian embryogenesis. The exprimental animal to be used is the preimplantation rabbit embryo. During the current project period, the research will involve three aspects of nucleic acid and protein metabolism in these embryos: (1) messenger RNA populations in rabbit oocytes and preimplantation embryos; (2) messenger RNA modifications and peptide chain initiation; and (3) DNA fragments identified on the surface of the trophoblast cells. Methods to be used involve the preparation of undegraded total cell and cytoplasmic RNA from oocytes and embryos, purifying it by cesium chloride centrifugation, and using it either in RNA-in-excess hybridizations with sonicated, iodinated single-copy rabbit DNA or in the preparation of cDNA, using the reverse transcriptase of the avian myeloblastosis virus. Other methods to be employed will involve the cell-free translation of messenger RNAs from rabbit blastocysts, and the assay of ternary complex formation between methionyl initiator transfer RNA, GTP, and the protein initiation factor eIF-2 from blastocysts. Further studies will involve the electron microscopy of embryonic and other tissues after staining with a platinum-thymine complex, the use of specific anti-DNA antibodies, and laser microbeam analysis of DNA content of trophoblast cells.
