Five of the most variable loci detected in human DNA by hybridization with DNA fingerprint probes have been cloned and characterized. Each locus consists of a tandem-repetitive minisatellite, with repeat units ranging in length from 9 to 45 base pairs depending on the locus. All of these cloned minisatellites act as locus-specific hybridization probes, and detect extremely variable Mendelian loci with heterozygosities ranging from 90 to 99%. These five hypervariable loci, together with a previously-isolated minisatellite designated p lambda g3, are dispersed over four autosomes (chromosomes 1, 5, 7 and 12). Syntenic pairs on chromosomes 1 and 7 show no detectable pair-wise linkage, and thus these hypervariable loci show no evidence of clustering within the genome and should provide valuable markers for mapping inherited disease. The locus-specific minisatellites act as very sensitive hybridization probes, and can be pooled to detect several hypervariable loci simultaneously. The applications of these probes in individual identification, paternity testing and analysis of cell chimaerism are discussed, and are illustrated by an analysis of forensic specimens from two victims who had been sexually assaulted and murdered.

The KE family is a large three-generation pedigree in which half the members are affected with a severe speech and language disorder that is transmitted as an autosomal dominant monogenic trait. In previously published work, we localized the gene responsible (SPCH1) to a 5.6-cM region of 7q31 between D7S2459 and D7S643. In the present study, we have employed bioinformatic analyses to assemble a detailed BAC-/PAC-based sequence map of this interval, containing 152 sequence tagged sites (STSs), 20 known genes, and >7.75 Mb of completed genomic sequence. We screened the affected chromosome 7 from the KE family with 120 of these STSs (average spacing <100 kb), but we did not detect any evidence of a microdeletion. Novel polymorphic markers were generated from the sequence and were used to further localize critical recombination breakpoints in the KE family. This allowed refinement of the SPCH1 interval to a region between new markers 013A and 330B, containing approximately 6.1 Mb of completed sequence. In addition, we have studied two unrelated patients with a similar speech and language disorder, who have de novo translocations involving 7q31. Fluorescence in situ hybridization analyses with BACs/PACs from the sequence map localized the t(5;7)(q22;q31.2) breakpoint in the first patient (CS) to a single clone within the newly refined SPCH1 interval. This clone contains the CAGH44 gene, which encodes a brain-expressed protein containing a large polyglutamine stretch. However, we found that the t(2;7)(p23;q31.3) breakpoint in the second patient (BRD) resides within a BAC clone mapping >3.7 Mb distal to this, outside the current SPCH1 critical interval. Finally, we investigated the CAGH44 gene in affected individuals of the KE family, but we found no mutations in the currently known coding sequence. These studies represent further steps toward the isolation of the first gene to be implicated in the development of speech and language.

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Recent reports on transfection of mouse cells with DNA from the established human urinary bladder cancer cell lines T24, J82 and EJ (MGH-U1), and the presence of an identical genetic modification in T24 and EJ cells have led us to examine the identity of these and other cultures of urothelial origin. By the criteria of HLA-A-B-C typing 7 and isozyme analysis, we conclude that EJ (MGH-U1) and some cultures of J82 are in fact T24 cells. However, five other bladder cancer cell lines, J82 (CO'T), RT4, RT112, TCCSuP and SCaBER, are clearly distinct from T24 by HLA typing (ref. 7) and/or isozyme patterns.

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