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A general strategy for selecting insertion mutations in mice has been devised. Constructs lacking a promoter and including a [3-galactosidase gene, or a reporter gene encoding a protein with both [3-galactosidase and neomycin phosphotransferase activity, were designed so that activation of the reporter gene depends on its insertion within an active transcription unit. Such insertion events create a mutation in the tagged gene and allow its expression to be followed by [3-galactosidase activity. Introduction of promoter trap constructs into embryonic stem (ES) cells by electroporation or retroviral infection has led to the derivation of transgenic lines that show a variety of 13-galactosidase expression patterns. Intercrossing of heterozygotes from 24 strains that express [3-galactosidase identified 9 strains in which homozygosity leads to an embryonic lethality. Because no overt phenotype was detected in the remaining strains, these results suggest that a substantial proportion of mammalian genes identified by this approach are not essential for development.