CYP2F1, which encodes a P450 enzyme capable of metabolizing several mono-oxygenase substrates with the highest activity toward ethoxycoumarin, has been mapped to human chromosome 19 by somatic cell hybrid studies. The CYP2A and CYP2B subfamilies are known to lie within 350 kb of each other on chromosome 19. To determine the locations of CYP2F1 with respect to CYP2A and CYP2B, multipoint linkage analysis and pulsed-field gel electrophoresis were performed. No recombinants were found between CYP2F1 and CYP2B in more than 50 meioses, and one recombinant was found between CYP2F1 and CYP2A (50 meioses). Pulsed-field gel electrophoresis showed the three loci to lie within a 240-kb region. Multipoint analysis of the haplotyped CYP cluster with four other chromosome 19 loci yielded the order D19S7-D19S9-APOC2-D19S8-CYP, although four other orders could not be excluded. The odds were 4.4 x 10(3) against the order proposed by the HGM10 consortium, D19S7-D19S9-CYP-D19S8-APOC2.