Synthesis of phospholipid during cleavage, compaction and blastocyst formation of the preimplantation mouse embryo was investigated using [methyl-3H]-choline as a specific precursor. The only choline-containing lipids found to incorporate label were phosphatidylcholine and lysolecithin. [Methyl-3H]choline incorporation into lipid was detectable at the 2-cell stage and increased 9-13-fold (on a per embryo basis) during the 8-cell stage and subsequent compaction of the morula. Incorporation of choline was also elevated in the blastocyst but could not be compared accurately with the rates observed in earlier embryos due to uncertainty about the size of the endogenous choline pool at this stage. Choline kinase (assayed in vitro) was detectable at every stage, its activity increased during development and paralleled (qualitatively) the extent of phosphocholine formation in intact embryos. Phospholipid turnover and choline base exchange did not contribute significantly to [methyl-3H]choline incorporation into lipid, which is hence judged to represent denovo synthesis of phospholipid via the Kennedy pathway. Mouse embryo lipids exhibit several features which may be characteristic of immature cells and which could influence the properties of their membranes. These include the absence of detectable sphingomyelin synthesis and the presence of demonstrable deacylation and turnover of phosphatidylcholine.