The coding region of Hand1 from a 600-bp cDNA initiating at the ATG and terminating at the stop codon followed by an EGFP expression cassette and a PGKNeo cassette flanked by loxP sites were inserted in frame at the position of the initiation codon. EGFP epifluorescence is not detected in E9.5 embryos, however EGFP mRNA is detected in a pattern identical to Hand1. qRT-PCR indicates that heterozygotes express about 90% of wild-type levels while homozygotes express at 50% that of wild-type, indicating that this is a hypomoprhic allele. In situ analysis indicates that expression is not uniformly reduced throughout the embryo.