The isolation of several distinct transcriptional regulatory factors required for myogenic determination of pluripotent stem cells has been recently reported. The molecular cloning and characterization of this set of structurally and functionally related regulatory genes allows a molecular analysis of events involved in the terminal differentiation of muscle cells. in vertebrates, the myo genic lineage derives from mesoderm which becomes segregated from the ectoderm and endoderm at the gastrula stage of embryonic development. Little is known about germ line specification in vertebrate embryos at the molecular level, but it is reasonable to anticipate that regulatory factors participating in mesoderm development and myogenic determination early in embryogenesis might be related to those involved in the later stages of muscle cell differentiation. We have recently isolated several MyoD1-related cDNAs that are present in Xenopus embryos prior to somite formation and muscle cell differentiation; these clones were designated XLMF, for Xenopus laevis myogenic factor. The purpose of this study is to further characterize these cDNA sequences, to examine their role in signaling the specification of myogenic cell lineages in developing frog embryos and the regulation of their expression, and to determine whether additional related genes are expressed early in development.