Haplotypes without children: PCR applied to close loci on individual human sperm

Indirect evidence of interindividual variation of the recombination fraction is considerable, but as yet direct evidence is lacking. Such interindividual variation could explain the widely observed gametic association in the major histocompatibility complex (MHC) region. A possible approach to this is the application of the polymerase chain reaction to closely linked markers on individual sperm. The feasibility of this approach is investigated here. Three loci in the human MHC (chromosome 6) were used in this study. Two are polymorphisms identifiable by oligonucleotide dot blotting, and one is a microsatellite polymorphism. The genetic distances between these loci are 0.5 and 1 centi-Morgan. We were able to double-type most cases for the two markers HLA-DPB and HLA-DRB in single haploid cells. The rate of double typings is comparable with that expected from the hypothesis of independent failures at two loci. However, this rate of failure remains too great to allow an analysis of segregation because allele specific failure cannot be ruled out. All double and triple typings (implicating the microsatellite) were perfectly correlated with each other; thus no recombinations were identified. With such close markers recombination would have been surprising and may indicate an inherent accuracy of positive haplotyping.