Biomedical subjects

Heinz-Ulrich G Weier

Publications and source records attributed to Heinz-Ulrich G Weier.

6 recordsLinked to original sources

PubMed2006

Chromosome-specific DNA repeat probes.

In research as well as in clinical applications, fluorescence in situ hybridization (FISH) has gained increasing popularity as a highly sensitive technique to study cytogenetic changes. Today, hundreds of commercially available DNA probes serve the basic needs of the biomedical research community. Widespread applications, however, are often limited by the lack of appropriately labeled, specific nucleic acid probes. We describe two approaches for an expeditious preparation of chromosome-specific DNAs and the subsequent probe labeling with reporter molecules of choice. The described techniques allow the preparation of highly specific DNA repeat probes suitable for enumeration of chromosomes in interphase cell nuclei or tissue sections. In addition, there is no need for chromosome enrichment by flow cytometry and sorting or molecular cloning. Our PCR-based method uses either bacterial artificial chromosomes or human genomic DNA as templates with alpha-satellite-specific primers. Here we demonstrate the production of fluorochrome-labeled DNA repeat probes specific for human chromosomes 17 and 18 in just a few days without the need for highly specialized equipment and without the limitation to only a few fluorochrome labels.

Chromosomes, Artificial, Bacterial↗

PubMed2006

Quantitative DNA fiber mapping in genome research and construction of physical maps.

Efforts to prepare a first draft of the human DNA genomic sequence forced multidisciplinary teams of researchers to face unique challenges. At the same time, these unprecedented obstacles stimulated the development of many highly innovative approaches to biomedical problem solving, robotics, and bioinformatics. High-resolution physical maps are required for ordering individual segments of information for the construction of a comprehensive map of the entire genome. This chapter describes a novel way to identify, delineate, and characterize selected, often small DNA sequences along a larger piece of the human genome. The technology is based on immobilization of high molecular weight DNA molecules on a solid substrate (such as a glass slide) followed by uniform stretching of the DNA molecule by the force of a receding meniscus. The hydrodynamic force stretches the DNA molecules homogeneously to approximately 2.3 kb/microm, so that distances measured after probe binding in microm can be converted directly into kb distances. Out of a large number of applications, this article focuses on mapping of genomic sequences relative to one another, the assembly of physical maps with near kb resolution, and, finally, quality control during physical map assembly and sequencing.

Chromosomes, Artificial↗

PubMed2005

Human cytotrophoblasts acquire aneuploidies as they differentiate to an invasive phenotype.

Through an unusual differentiation process, human trophoblast progenitors (cytotrophoblasts) give rise to tumor-like cells that invade the uterus. By an unknown mechanism, invasive cytotrophoblasts exhibit permanent cell cycle withdrawal. Here, we report molecular cytogenetic data showing that approximately 20 to 60% of these interphase cells had acquired aneusomies involving chromosomes X, Y, or 16. The incidence positively correlated with gestational age and differentiation to an invasive phenotype. Scoring 12 chromosomes in flow-sorted cytotrophoblasts showed that more than 95% of the cells were hyperdiploid. Thus, aneuploidy appears to be an important component of normal placentation, perhaps limiting the proliferative and invasive potential of cytotrophoblasts within the uterus.

Aneuploidy↗

PubMed2005

Independence of replisomes in Escherichia coli chromosomal replication.

In Escherichia coli DNA replication is carried out by the coordinated action of the proteins within a replisome. After replication initiation, the two bidirectionally oriented replisomes from a single origin are colocalized into higher-order structures termed replication factories. The factory model postulated that the two replisomes are also functionally coupled. We tested this hypothesis by using DNA combing and whole-genome microarrays. Nascent DNA surrounding oriC in single, combed chromosomes showed instead that one replisome, usually the leftward one, was significantly ahead of the other 70% of the time. We next used microarrays to follow replication throughout the genome by measuring DNA copy number. We found in multiple E. coli strains that the replisomes are independent, with the leftward replisome ahead of the rightward one. The size of the bias was strain-specific, varying from 50 to 130 kb in the array results. When we artificially blocked one replisome, the other continued unabated, again demonstrating independence. We suggest an improved version of the factory model that retains the advantages of threading DNA through colocalized replisomes at about equal rates, but allows the cell flexibility to overcome obstacles encountered during elongation.

Chromosomes↗

PubMed2005

Aneuploidy involving chromosome 1 in failed-fertilized human oocytes is unrelated to maternal age.

PURPOSE: To study whether maternal meiotic errors in failed-fertilized oocytes involving chromosome 1 occur at frequencies similar to those involving other autosomes, and whether their frequency is affected by maternal age. METHODS: Using fluorescence in situ hybridization (FISH), frequencies of aneusomy and chromatid pre-division involving chromosomes 1, 16, 18, and 21 were determined for 273 failed-fertilized oocytes. RESULTS: The aneuploidy rate for chromosome 1 was 15.8%, and was neither age-dependent nor significantly different from that for chromosomes 16, 18 or 21. Only chromosome 16 exhibited an age-dependent increase in aneusomy rates. The frequency of chromatid pre-division was lower for chromosome 1 than for chromosome 18 (11.9% vs. 25.4%; p = 0.01), but not different from that for chromosomes 16 or 21. CONCLUSION: Aneuploidy involving chromosome 1 in failed-fertilized oocytes is unrelated to maternal age and occurs at a frequency similar to that for chromosomes 16, 18, and 21.

Aneuploidy↗

PubMed2002

FISH in cancer diagnosis and prognostication: from cause to course of disease.

The last 20 years have witnessed an astounding evolution of cytogenetic approaches to cancer diagnosis and prognostication. Molecular techniques and, in particular, nonisotopically-labeled nucleic acid probes and fluorescence in situ hybridization (FISH)-based techniques have replaced the costly and potentially dangerous radioactive techniques used in research and the clinical detection of genetic alterations in tumor cells. Fluorescent DNA probes also enabled the screening for very subtle chromosomal changes. Clinical laboratories now choose from a growing number of FISH-based cytogenetic tests to support physician's diagnoses of the causes and the course of a disease. Depending on the specimen, state-of-the-art FISH techniques allow the localization and scoring of 10-24 different targets and overcome previous problems associated with target colocalization and detection system bandwidth. FISH-based analyses have been applied very successfully to the analysis of single cells and have demonstrated the existence of cell clones of different chromosomal make-up within human tumors. This information provides disease-specific information to the attending physician and should enable the design of patient-specific protocols for disease intervention.

Chromosome Aberrations↗