Chromosome jumping from D4S10 (G8) toward the Huntington disease gene.

Abstract

The gene for Huntington disease (HD) has been localized to the distal portion of the short arm of human chromosome 4 by linkage analysis. Currently, the two closest DNA markers are D4S1O (G8), located "p3 centimorgans centromeric to HD, and D4S43 (C4H), positioned 0-1.5 centimorgans from HD. In an effort to move closer to the HD gene, with the eventual goal of identifying the gene itself, we have applied the technique of chromosome jumping to this region. A 200-kilobase jumping library has been constructed, and ajump from D4S1O has been obtained and its approximate distance verified by pulsed field gel electrophoresis. Two restriction fragment length polymorphisms have been identified at the jump locus, which is denoted D4S81. Linkage analysis of previously identified recombinants between D4S1O and HD or D4S1O and D4S43 shows that in two of five events the jump has crossed the recombination points. This unequivocally orients D4S1O and D4S81 on the chromosome, provides additional markers for HD, and suggests that recombination frequency in this region of chromosome 4 may be increased, so that the physical distance from D4S1O to HD may not be as large as originally suspected.A large number of human disorders are caused by mutations in single genes whose normal function, despite intensive investigation, is unknown. Recently, it has become possible to attempt the cloning of such genes by a process called " reverse genetics," in which the gene is identified primarily by its map position in the genome rather than by direct knowledge of its protein product (1, 2). Huntington disease (HD) is an autosomal dominant neurodegenerative condition with a wide range in age of onset (3, 4). The motor manifestations include incoordination and involuntary movements (chorea). Psychiatric features of forgetfulness, personality changes, depression, and eventual dementia are characteristic. Anatomically, the most pronounced changes are neuronal loss in the basal ganglia of the brain, especially the caudate nucleus; thus far, however, no specific protein abnormality has been found.HD has been found to be tightly linked (5) to the DNA marker D4SJ0 (G8), which has been mapped by in situ hybridization (6) and somatic cell genetics (7) (13) and independently by Poutska and Lehrach (14), depends on the circularization of very large DNA fragments, followed by cloning of the junction fragments of these circles, which bring together DNA sequences that were originally located a considerable distance apart in the genome. We have previously demonstrated the successful application of this technique to a jump of 100 kb in the cystic fibrosis region (15). In this report, we describe the construction of a 200-kb jumping library and its successful utilization to move closer to the HD gene from D4S10.MATERIALS AND METHODS Cell Line. HHW661 is a somatic cell hybrid carrying a selectable human t(4;5) (p15.1;pl5.1) translocation chromosome as its only human DNA (16

New Insights Into the Clinical Features, Pathogenesis and Molecular Genetics of Huntington Disease

“…Firstly, the analysis of informative crossover events in HD pedigrees and secondly, the assessment of linkage disequilibrium of DNA markers spanning the entire 6 megabase region with the disease, have been included. These studies have initiated an extensive search for new polymorphic DNA markers from the HD region distal to D4S10 (130,(133)(134)(135)(136)(137)(138)(139)(140)(141) and have also facilitated the construction of detailed genetic (142) and physical maps (143,144) of the 4p16.3 region (Fig. 7).…”

A chromosome jump crosses a translocation breakpoint in the von recklinghausen neurofibromatosis region

“…One can usually proceed further along the chromosome in fewer steps, and can skip over unclonable regions that would stop traditional walking. General jumping libraries can be constructed from total genomic DNA, such as used here, or from hybrids for specialized purposes (Richards et al, 1988). T h e library used here was created by cloning into the EcoRI site of the phage vector; other enzymes can be used in construction to compensate for any cloning bias associated with a particular enzyme.…”

Chromosomal jumping from the DXS165 locus allows molecular characterization of four microdeletions and a de novo chromosome X/13 translocation associated with choroideremia.

“…3). DISCUSSION So far, there is only a limited number of reports dealing with the successful application of chromosome-jumping techniques (21,(23)(24)(25)27). In this study, we made use of a 100-kb human chromosomal-jumping library that had been used previously to study the chromosomal region carrying the cystic fibrosis locus (21) and the Duchenne gene (24).…”

“…Chromosome jumping is based on the circularization of long genomic DNA fragments and subsequent cloning of the junction fragments of these circles (20)(21)(22)(23)(24)(25)(26)(27). In this way, it is possible to circumvent insert-size constraints of conventional cloning vectors, which limit the distance that can be covered by individual chromosome walking steps.…”