The majority of exon 2, which encodes the basic helix-loop-helix domain, was replaced with a cassette containing both IRES-lacZ and neo genes. Transcript was undetected by RT-PCR analyses of homozygous mutant embryos. Weak expression of beta-galactosidase, putatively due to the inefficiency of the IRES, showed the pattern of expression to recapitulate that of the endogenous gene.