The CBFβ subunit is the non-DNA-binding subunit of the heterodimeric core-binding factor (CBF). CBFβ associates with DNA-binding CBFα subunits and increases their affinity for DNA. Genes encoding the CBFβ subunit (CBFB) and one of the CBFα subunits (CBFA2, otherwise known as AML1) are the most frequent targets of chromosomal translocations in acute leukemias in humans. We and others previously demonstrated that homozygous disruption of the mouse Cbfa2 (AML1) gene results in embryonic lethality at midgestation due to hemorrhaging in the central nervous system and blocks fetal liver hematopoiesis. Here we demonstrate that homozygous mutation of the Cbfb gene results in the same phenotype. Our results demonstrate that the CBFβ subunit is required for CBFα2 function in vivo.