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G D Stewart

Publications and source records attributed to G D Stewart.

At least 19 recordsLinked to original sources
PubMed1992

Sequence-tagged sites (STSs) for a set of mapped markers on chromosome 21.

Sequence tagged sites (STSs) have been proposed as a "common language" for comparing physical and genetic maps of the human genome produced by a variety of techniques. We have produced 44 STSs from 38 mapped loci on human chromosome 21. The STSs represent most of the loci designated as genetic reference or ordered physical framework markers, along with a number of others chosen to span all regions of 21q. Of the STSs, 12 are from gene segments, including 4 from exons of the APP gene encoding the amyloid beta protein precursor, and 32 mark anonymous DNA loci. These STSs make each of the corresponding loci readily accessible to the research community without the need for exchange of clones. These sites also represent multiple start points for the isolation of YAC clones that should permit overlapping the entire chromosome 21 long arm as cloned DNA.

Animals↗
PubMed1992

A genetic linkage map of human chromosome 21: analysis of recombination as a function of sex and age.

A genetic linkage map of human chromosome 21 has been constructed using 22 anonymous DNA markers and five complementary DNAs (cDNAs) encoding the amyloid beta protein precursor (APP), superoxide dismutase 1 (SOD1), the ets-2 proto-oncogene (ETS2), the estrogen inducible breast cancer locus (BCEI), and the leukocyte antigen, CD18 (CD18). Segregation of RFLPs detected by these DNA markers was traced in the Venezuelan Reference Pedigree (VRP). A comprehensive genetic linkage map consisting of the 27 DNA markers spans 102 cM on the long arm of chromosome 21. We have confirmed our initial findings of a dramatically increased rate of recombination at the telomere in both females and males and of significantly higher recombination in females in the pericentromeric region. By comparing patterns of recombination in specific regions of chromosome 21 with regard to both parental sex and age, we have now identified a statistically significant downward trend in the frequency of crossovers in the most telomeric portion of chromosome 21 with increasing maternal age. A less significant decrease in recombination with increasing maternal age was observed in the pericentromeric region of the chromosome. These results may help in ultimately understanding the physical relationship between recombination and nondisjunction in the occurrence of trisomy 21.

Adolescent↗
PubMed1991

Plasmids for recombination-based screening.

To facilitate recombination-based screening, we constructed the ColE1-based plasmid, pi G4, that confers chloramphenicol resistance, contains a polylinker with multiple unique restriction enzyme recognition sequences, and contains the genetic marker, supF. To facilitate recombination-based screening followed by rapid DNA sequencing, we inserted the selectable marker, supF, into each of 20 high-copy-number (hcn) pUC-derived NoC plasmids that were designed for multiplex DNA sequencing. To facilitate recombination-based screening of common cDNA libraries that often contain ColE1 sequences, we constructed a supF-carrying plasmid whose replication was driven from an R6K replicon that does not share sequence homology with ColE1. Furthermore, we incorporated a useful polylinker and increased the copy number of this plasmid to create the 4.4-kb hcn plasmid, pMAD1. Thus, these plasmids allow: (1) background-free transformation of cells by a supF plasmid carrying an antibiotic-resistance marker; (2) simultaneous performance of the recombination-based assay and DNA sequencing; and (3) screening bacteriophage cDNA libraries that contain ColE1 sequences by recombination with a supF plasmid that is not homologous to ColE1 derivatives.

Bacteriophage lambda↗
PubMed1991

DNA sequences surrounding the centromere of chromosome 21.

The mapping and sequencing of two clones that surround the centromere of chromosome 21 are presented. These clones specify the most proximal known low-order repeat on 21p (p21-7D) and the most proximal known single-copy sequence on 21q (pUT-B37 at locus D21S120).

Base Sequence↗
PubMed1990

Recombination-based screening for genes on chromosome 21.

In previous work, a reasonably large number of genes was found to be on chromosome 21. This frequency demands the creation of new techniques to investigate the transcription of this small human autosome, and to relate these findings to the phenotype of Down syndrome. Here we describe the elaboration of new vectors and hosts, which in conjunction with flow-sorted cosmid libraries of chromosome 21 and genic (cDNA) libraries of relevant human tissues, will permit us to examine transcription of chromosome 21.

Chromosome Mapping↗
PubMed1990

Identification of Campylobacter pylori by endoscopic brush cytology.

To investigate the value of Papanicolaou-stained endoscopic brush samples in the diagnosis of Campylobacter pylori infection of the upper gastrointestinal tract, 138 brush and biopsy samples from the esophagus, stomach, and duodenum, taken concomitantly, were reviewed retrospectively. In 35 cases, Campylobacter-like organisms (CLOs) were found in both cytology and biopsy samples. In 15 cases, CLOs were seen in biopsy material only, and in 8 cases, CLOs were found in cytology material only. CLOs were found in 49% of the gastric specimens and 33% of the Barrett's esophagus specimens by histologic or cytologic examination or by both methods. CLOs were found by at least one method in 64% of the gastric samples with active gastritis 40% with borderline gastritis, 15% without gastritis, and in 64% with adenocarcinoma. Cytologic examination of endoscopic brush samples is a valuable technique for the diagnosis of gastric Campylobacter infections and can be performed easily in cytopathology laboratories.

Biopsy↗
PubMed1990

The murine situs inversus viscerum (iv) gene responsible for visceral asymmetry is linked tightly to the Igh-C cluster on chromosome 12.

The iv gene controls left-right determination during murine organogenesis. To map this gene, we analyzed backcross progeny produced by mating (C57BL/6J X MEV/Ty)F1-iv/+heterozygotes to C57BL/6J-iv homozygotes. Hybridization of a murine ecotropic virus probe and several homeotic box gene probes coupled with analysis of dominant visible markers enabled us to exclude the iv locus from much of the mouse genome. Spurred by a recent report that mapped the iv gene to mouse chromosome 12 which was not excluded by our previous work, we used the polymerase chain reaction on our larger cohort to determine that the iv gene is indeed linked tightly to the Igh-C locus on this chromosome: we observed 0/156 recombinants between the iv and Igh-C loci. Combining data from the two studies demonstrates that the murine iv gene is close (1/201 recombinants) to the Igh-C cluster on chromosome 12.

Animals↗
PubMed1989

Characterization of an unusual and complex chromosome 21 rearrangement using somatic cell genetics and cloned DNA probes.

In a previous case of a newborn infant with typical Down syndrome, chromosome analysis indicated the presence of an unusual and complex translocation of chromosome 21. The patient's cells contained one normal chromosome 21 and a rearranged, F group-sized submetacentric chromosome. This abnormal chromosome appeared to involve duplication of the distal portion of 21q with translocation to the short arm, and a deletion of C-band-positive centromeric heterochromatin. Using linearly ordered cloned DNA probes, we report the detailed molecular examination of this abnormal chromosome, which has been isolated on a hamster background in a hybrid cell line. Both short arm and pericentromeric sequences are present on this chromosome, as well as distal 21q sequences. However, a substantial portion of proximal 21q is deleted. The distal boundary of this deleted section can be pinpointed within the region between two loci (D21S8 and D21S54), a distance of about 5,000 kb. This study illustrates the power of using precisely mapped, linearly ordered DNA probes to characterize this type of rearrangement. In addition, this hybrid cell line can also be used as a member of a mapping panel to map DNA sequences regionally on chromosome 21.

Animals↗
PubMed1988

Isolation of DNA sequences on human chromosome 21 by application of a recombination-based assay to DNA from flow-sorted chromosomes.

By merging two efficient technologies, bivariate flow sorting of human metaphase chromosomes and a recombination-based assay for sequence complexity, we isolated 28 cloned DNA segments homologous to loci on human chromosome 21. Subregional mapping of these DNA segments with a somatic cell hybrid panel showed that 26 of the 28 cloned DNA sequences are distributed along the long arm of chromosome 21, while the other 2 hybridize with sequences on the short arm of both chromosome 21 and other chromosomes. This new collection of probes homologous to chromosome 21 should facilitate molecular analyses of trisomy 21 by providing DNA probes for the linkage map of chromosome 21, for studies of nondisjunction, for chromosome walking in clinically relevant subregions of chromosome 21, and for the isolation of genes on chromosome 21 following the screening of cDNA libraries.

Animals↗
PubMed1988

Genetic linkage map of human chromosome 21.

Two of the most common disorders affecting the human nervous system, Down syndrome and Alzheimer's disease, involve genes residing on human chromosome 21. A genetic linkage map of human chromosome 21 has been constructed using 13 anonymous DNA markers and cDNAs encoding the genes for superoxide dismutase 1 (SOD1) and the precursor of Alzheimer's amyloid beta peptide (APP). Segregation of restriction fragment length polymorphisms (RFLPs) for these genes and DNA markers was traced in a large Venezuelan kindred established as a "reference" pedigree for human linkage analysis. The 15 loci form a single linkage group spanning 81 cM on the long arm of chromosome 21, with a markedly increased frequency of recombination occurring toward the telomere. Consequently, 40% of the genetic length of the long arm corresponds to less than 10% of its cytogenetic length, represented by the terminal half of 21q22.3. Females displayed greater recombination than males throughout the linkage group, with the difference being most striking for markers just below the centromere. Definition of the linkage relationships for these chromosome 21 markers will help refine the map position of the familial Alzheimer's disease gene and facilitate investigation of the role of recombination in nondisjunction associated with Down syndrome.

Chromosome Mapping↗
PubMed1988

Trisomy 21 (Down syndrome): studying nondisjunction and meiotic recombination by using cytogenetic and molecular polymorphisms that span chromosome 21.

By combining molecular and cytogenetic techniques, we demonstrated the feasibility and desirability of a comprehensive approach to analysis of nondisjunction for chromosome 21. We analyzed the parental origin and stage of meiotic errors resulting in trisomy 21 in each of five families by successfully using cytogenetic heteromorphisms and DNA polymorphisms. The 16 DNA fragments used to detect polymorphisms spanned the length of the long arm and detected recombinational events on nondisjoined chromosomes in both maternal meiosis I and maternal meiosis II errors. The meiotic stage at which errors occurred was determined by sandwiching the centromere between cytogenetic heteromorphisms on 21p and an informative haplotype constructed using two polymorphic DNA probes that map to 21q just below the centromere. This study illustrates the necessity of combining cytogenetic polymorphisms on 21p with DNA polymorphisms spanning 21q to determine (1) the source and stage of meiotic errors that lead to trisomy 21 and (2) whether an association exists between nondisjunction and meiotic recombination.

Chromosome Banding↗
PubMed1988

Molecular and cytogenetic characterization of a de novo t(5p;21q) in a patient previously diagnosed as monosomy 21.

Genomic single-copy DNA fragments were used to characterize an undetected chromosome translocation in an individual whose metaphase chromosome analysis revealed apparent monosomy 21. Eight RFLPs detected by six probes were used to identify homologous sequences from chromosome 21 in DNA digests from the proband and her parents. These family studies showed that the proband was disomic for the distal region of 21q. Reverse banding and in situ hybridization of chromosome 21-specific probes to metaphase chromosomes from the proband revealed a de novo translocation with breakpoints at 5p13 or 14 and 21q11 or 21. In situ hybridization permitted orientation of the translocated portion of chromosome 21 on the derivative chromosome 5 and, in conjunction with molecular analysis and previous mapping studies, refined the physical map for the long arm of chromosome 21.

Child↗
PubMed1987

Isolation of polymorphic DNA fragments from human chromosome 4.

We have identified and characterized 40 DNA probes detecting restriction fragment length polymorphism (RFLP) on human chromosome 4. Single copy human clones were isolated from a bacteriophage library enriched for chromosome 4 sequences. Each clone was hybridized to somatic cell hybrid DNAs for verification of its species and chromosomal origin and for regional localization. Sequences specific for chromosome 4 were tested for their ability to detect RFLPs in human DNA and their potential utility as genetic markers was assessed. Approximately 263,000 base pairs or 0.13% of the chromosome was screened for sequence variation. The estimate of heterozygosity calculated from this large body of data, H = 0.0021, indicates that the degree of sequence variation on chromosome 4 is comparable to other autosomes. The characterization of these 40 markers has tripled the number of polymorphic loci available for linkage studies on chromosome 4, making it feasible to begin construction of a detailed linkage map that will span the entire chromosome.

Animals↗