The two human genes FMO2 and FMO5 are shown to be located on the long arm of chromosome 1 (1q) through the screening of human-rodent somatic cell hybrids by PCR. Fifteen YAC clones shown to contain one or more FMO genes within their inserts were isolated from an ICRF YAC human genomic DNA library. Two clones were found to contain FMO1, FMO2, FMO3 and FMO4 within their inserts. All clones containing one or more of FMO1, FMO2, FMO3 or FMO4 were found by fluorescence in-situ hybridization (FISH) to human metaphase chromosomes to hybridize at 1q23-24. Four clones were found to contain FMO5: in each case, none of the other known FMO genes were also present. FISH to human metaphase chromosomes with these four clones revealed hybridization at 1q21. Subjecting three of the YAC clones, two containing FMO5 and the other FMO1, FMO2, FMO3 and FMO4, to pulsed field gel electrophoresis, allowed estimation of their insert sizes. Screening of an ICRF chromosome 1-specific cosmid library resulted in the isolation of five cosmid clones containing inserts with apparent homology to FMO sequences. Southern hybridization and FISH analysis, however, indicated that the clone inserts did not contain FMO genes. Rather, four inserts appeared to originate from heterochromatic regions and the other from 15q11-13.