A method for obtaining a pure population of t6/t6 mouse embryos prior to developmental arrest

Developmental delay, as the result of ovariectomy, causes mouse blastocyst embryos obtained from +/t6 inter se matings to separate into two distinct populations when placed into outgrowth medium. One population remains as free floating embryos for a significantly longer period of time than the other population. Based upon their phenotypic expression following attachment and outgrowth, the former population was considered to be composed entirely of t6/t6 embryos and the latter, to be composed of +/+ and +/t6 embryos (Nadijcka et al., '81). In the present study, two-dimensional gel electrophoresis identified the embryos which were delayed in attachment as t6/t6 embryos since they synthesize only p63/6.9a, a product of the Tcp-1a locus which is unique to t-haplotypes. The early attaching embryos, assumed to be +/+ and +/t6, synthesize both p63/6.9a and b. The p63/6.9b protein is coded for by Tcp-1b on the wild-type homologous chromosome. Control +/+ blastocyst embryos synthesize only p63/6.9b. The data show that t6/t6 embryos can be identified prior to their lethal period and, thus, subjected to comparative studies to determine the cause of their lethality. Developmental delay is the first method established to enable one to unambiguously identify t6/t6 embryos prior to developmental arrest.