Mary Lou Orcutt

Localization of the von Hippel-Lindau disease gene to a small region of chromosome 3

We studied 25 families with von Hippel-Lindau disease (VHL) to locate VHL more precisely on chromosome 3. We found that VHL was linked to RAF1, confirming previous observations, and to two polymorphic DNA markers, D3S18 and D3S191. Multipoint linkage analysis indicated that the most likely location for VHL was in the interval between RAF1 and D3S18. D3S18 was located at 3p26. Genetic heterogeneity was not detected in this panel of von Hippel-Lindau disease families. The polymorphic markers RAF1, D3S18, and D3S191 should be useful in identifying asymptomatic gene carriers in VHL families and in guiding efforts at gene isolation.

Von Hippel-Lindau disease: identification of deletion mutations by pulsed-field gel electrophoresis

Von Hippel-Lindau disease (VHL) is an inherited multisystem neoplastic disorder. We prepared a 2.5-megabase (Mb) restriction map of the region surrounding the VHL gene and identified and characterized overlapping deletions in three unrelated patients affected with VHL. The smallest nested deletion (100 kb) was located within a 510-kb NruI fragment detected by 19–63′. The rearrangements detected will be useful in isolating and evaluating candidate cDNAs for the VHL gene. The detailed physical map will be useful in studying the organization and structure of genes in the VHL region.

Isolation and regional localization of a large collection (2,000) of single-copy DNA fragments on human chromosome 3 for mapping and cloning tumor suppressor genes

SummaryA collection of 2,000 lambda phage-carrying human single-copy inserts (> 700 bp) were isolated from two chromosome-3 flow-sorted libraries. The single-copy DNA fragments were first sorted into 3p and 3q locations and about 700 3p fragments were regionally mapped using a deletion mapping panel comprised of two humanhamster and two-human-mouse cell hybrids, each containing a chromosome 3 with different deletions in the short arm. The hybrids were extensively mapped with a set of standard 3p markers physically localized or ordered by linkage. The deletion mapping panel divided the short arm into five distinct subregions (A-E). The 3p fragments were distributed on 3p regions as follows: region A, 26%; B, 31%; C, 4%; D, 4% and E, 35%. We screened 300 single-copy DNA fragments from the distal part of 3p (regions A and B) with ten restriction endonucleases for their ability to detect restriction fragment length polymorphisms (RFLPs). Of these fragments 110 (36%) were found to detect useful RFLPs: 35% detected polymorphisms with frequency of heterozygosity of 40% or higher, and 25% with frequency of 30% or higher. All polymorphisms originated from single loci and most of them were of the base pair substitution type. These RFLP markers make it possible to construct a fine linkage map that will span the distal part of chromosome 3p and encompasses the von Hippel-Lindau disease locus. The large number of single-copy fragments (2,000) spaced every 100–150 kb on chromosome 3 will make a significant contribution to mapping and sequencing the entire chromosome 3. The 300 conserved chromosome 3 probes will increase the existing knowledge of man-mouse homologies.

Molecular and genetic characterization and physical mapping of 11 new markers detecting multiallele restriction fragment length polymorphisms on the short arm of human chromosome 3

SummaryGenetic markers with high degrees of polymorphisms are of vital importance in the construction of high resolution (2–4 cM) linkage maps of human chromosomes as specified in the short-term goals of the Human Genome Initiative. In this paper, we report on molecular and genetic characterization and physical localization of 11 new multiallele restriction fragment length polymorphism markers on human chromosome 3p. Ten of these represent three- and four-allele polymorphisms of the base substitution type probably at two adjacent restriction sites. One has been identified as a novel minisatellite sequence comprising a variable copy number tandem repeat array of a G/T-rich 79-bp sequence. This collection of multiallele polymorphic (PIC values: 0.40–0.60) markers should prove valuable and increase the resolution power of the available chromosome 3p genetic markers.