Preimplantation diagnosis of the delta F508 deletion causing cystic fibrosis is possible through in vitro fertilization, biopsy of a cleavage-stage embryo, and amplification of DNA from single embryonic cells. This approach should be equally applicable to other single-gene diseases in which the defect has been identified. Analysis of a series of pregnancies, however, will be required to assess the method adequately.

Choroideremia, an X-chromosome linked retinal dystrophy of unknown pathogenesis, causes progressive nightblindness and eventual central blindness in affected males by the third to fourth decade of life. Choroideremia has been mapped to Xql3-21 by tight linkage to restriction fragment length polymorphism loci. We have recently identified two families in which choroideremia is inherited with mental retardation and deafness. In family XL-62, an interstitial deletion in Xq21 is visible by cytogenetic analysis and two linked anonymous DNA markers, DXYS1 and DXS72, are deleted. In the second family, XL-45, an interstitial deletion was suspected on phenotypic grounds but could not be confirmed by high-resolution cytogenetic analysis. We used phenol-enhanced reassociation of 48,XXXX DNA in competition with excess XL-45 DNA to generate a library of cloned DNA enriched for sequences that might be deleted in XL-45. Two of the first 83 sequences characterized from the library were found to be deleted in probands from family XL-45 as well as from family XL-62. Isolation of these sequences proves that XL-45 does contain a submicroscopic deletion and provides a starting point for identifying overlapping genomic sequences that span the XL-45 deletion. Each overlapping sequence will be studied to identify exons from the choroideremia locus. these families, XL-62, an interstitial deletion within Xq21 was found cytogenetically and confirmed by Southern blot analysis of the probands' DNA by demonstrating the deletion of two RFLP loci, DXYS1 and DXS72, known to be tightly linked to choroideremia. In the second family, XL-45, an unequivocal interstitial deletion could not be demonstrated cytogenetically and no linked markers were found to be deleted (see below). However, similarities in phenotype between the affected males in the two families led us to hypothesize that the probands from the second family, XL-45, also had an interstitial deletion, albeit a much smaller one, responsible for the phenotype. To test this hypothesis, we used a phenol-enhanced reassociation technique (PERT) (9, 10), to enrich for DNA deleted in XL-45. With this approach, we have identified two DNA sequences that are deleted in the XL-45 probands, who lack an obvious cytogenetic deletion, as well as in the XL-62 probands, who do show a visible chromosomal deletion at Xq21. These probes are the first step toward isolating overlapping DNA sequences that span the submicroscopic deletion in XL-45, with the ultimate aim of identifying and isolating the choroideremia gene.

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