A locus in the human genome, not associated with any specific gene, has been found to be a site of restriction fragment length polymorphism. The polymorphism was found by hybridizing a 16-kilobase-pair segment of single-copy human DNA, selected from the human genome library cloned in phage XCH4A, to a Southern transfer of total human DNA digested with EcoRI. DNAs from a number of individuals from within Mormon pedigrees as well as random individuals have been examined. The locus is highly variable, with at least eight alleles present, homozygotes accounting for less than 25% of the individuals examined. The polymorphism appears to be the result of DNA rearrangements rather than base-pair substitutions or modifications. Examination of the DNA from seven members of a family revealed fragment lengths that are consistent with their inheritance as Mendelian alleles through three generations.Because of the scarcity of useful genetic marker loci, the human genetic linkage map has relatively few entries. For a marker locus to be useful in linkage studies, it must be the site of several variants (alleles). With several alleles occurring at reasonable frequencies in the population, parental individuals heterozygous at different marker loci will be common and cosegregation of markers can be scored among their progeny.Almost all polymorphic marker loci thus far identified in humans represent biochemical variations, such as isozymes or cell surface markers. Most of these loci do not have a sufficient number and frequency of alleles to be of general use in linkage studies in families, and many are not appropriate for prenatal diagnosis. A few biochemical and immunological loci having reasonably frequent variants within the human population have been described, however, and shown to be useful for linkage studies and for the resolution of genetic models (1).We have argued (2) that a large number of DNA sequence variations exist in the human population. Some of these should be detectable as variants in the lengths of the DNA fragments produced by a restriction enzyme (restriction fragment length polymorphism or RFLP). These loci will define arbitrary genetic sites, not necessarily associated with any specific genes, and should prove highly suitable as genetic markers. A linkage map of the human can be constructed by determining the pattern of cosegregation of these genetic markers in human pedigrees. A map based on several hundred such loci will have a sufficient marker density to allow the placement of any other locus with reasonable certainty (2, 3). RFLP markers specifically associated with the human ,3-and "y-globin loci have already been reported (4, 5). We will describe here an arbitrary locus showing a significant frequency of DNA sequence variation.In these experiments, the restriction fragments from the DNA of a locus are revealed by hybridization with a radiolabeledThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in...

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Flow cytometry and recombinant DNA techniques have been used to obtain reagents for a molecular analysis of the Prader-Willi syndrome (PWS). HindIll totaldigest libraries were prepared in X phage Charon 21A from flow-sorted inverted duplicated no. 15 human chromosomes and propagated on recombination-proficient (LE392) and recBC-, sbcB-(DB1257) bacteria. (2)(3)(4)(5)(6).The most common chromosome aberration in PWS patients is a deletion of band 15q11.2 (2); however, the size of these deletions is variable and presumably some are undetectable by using current cytogenetic techniques (see Fig. 1). Other aberrations at 15q11.2 occur in 2-5% of PWS patients (6-8), including translocations and duplications (both tandem and inverted), the latter termed inv dup(15) chromosomes.This multiplicity of chromosome rearrangements involving 15qll->13 in the PWS suggests that the proximal long arm of chromosome 15 contains DNA sequences, such as tandem or inverted repeats, which could predispose this segment to structural instability.To define this region of the human genome on a molecular level, several DNA segments from the proximal long arm of chromosome 15 were isolated from X phage Charon 21A libraries constructed from HindIII-digested DNA obtained from flow-sorted (9) inv dup(15) chromosomes. Because these inverted duplicated chromosomes contain two copies of all sequences within 15pter->15ql3, we were able to more easily isolate DNA segments from the proximal long arm of chromosome 15. Using different cloning and insert screening strategies we determined that many of these sequences are genetically unstable in a recombination-proficient bacterial strain. Electron micrographs of heteroduplexes showed that unstable clones contain inverted repeats, which suggests at least one possible mechanism for their instability. In total, we describe in this report ll different cloned DNA segments that map to 15qll-43; 4 of 8 tested are deleted in one of two PWS patients with a deletion of 15qll.2. These data suggest that there is molecular heterogeneity between 15qll deletions in different PWS patients and that the cytogenetic instability of band 15q11.2 might be explained in terms ofthe types ofDNA sequences it contains. MATERIALS AND METHODSCell Lines. The lymphoblastoid cell lines used were from a karyotypically normal (46,XY) individual (MD-li), two patients with the PWS and different-sized deletions in 15qll.2 (DON-5, and DON-10), and two individuals with inv dup(15) chromosomes of different size [ALD-6 (10) and ALD-24 (11)] (Fig. 1).Bacterial Strains. Strain DB1257 is related to strain DB1170 (12) and was constructed by P1 transduction of CES 200 (13) to kanamycin resistance using a lysate prepared on NK 5857 (Trp::TnS SupF 58). The partial genotype of CES 200 is recBC-, sbcB-. Strain LE392 has been described (14).Construction of the inv dup(15) Library. Charon 21A libraries were created from 50 ng of HindIII-digested DNA from 6 million flow-sorted, inv dup(15) chromosomes, isolated from cell line ALD-24. Chromosomes were...

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Use of a 4.5-kilobase-pair (kb) segment of single-copy DNA from a human genomic library as a hybridization probe of genomic human DNAs revealed allelic Taq I restriction fragments 10.6, 11.8, and 14.6 kb long. Among 12 unrelated individuals, all 6 males exhibited the 14.6-kb fragment in addition to one of the other fragments. Three of the females displayed 10.6- and 11.8-kb fragments, and the other three displayed only one fragment length; none had the 14.6-kb fragment. Hybridization of this probe to Taq I-digested DNAs from human-rodent hybrid cell lines (which have partial complements of human chromosomes) demonstrated segregation of the 14.6-kb fragment with the human Y chromosome and segregation of the 10.6- and 11.8-kb fragments with the human X chromosome. Furthermore, hybridization of this probe to Taq I-digested DNAs from 48 members of a single kindred revealed Y-linked inheritance of the 14.6-kb fragment and X-linked inheritance of the 10.6- and 11.8-kb fragments. These experiments demonstrate homology between single-copy sequences on the human X and Y chromosomes.