Showing papers by "Markus Grompe published in 1991"

A gene from the region of the human X inactivation centre is expressed exclusively from the inactive X chromosome

Abstract: X-chromosome inactivation results in the cis-limited dosage compensation of genes on one of the pair of X chromosomes in mammalian females. Although most X-linked genes are believed to be subject to inactivation, several are known to be expressed from both active and inactive X chromosomes. Here we describe an X-linked gene with a novel expression pattern--transcripts are detected only from the inactive X chromosome (Xi) and not from the active X chromosome (Xa). This gene, called XIST (for Xi-specific transcripts), is a candidate for a gene either involved in or uniquely influenced by the process of X inactivation.

Characterization of a murine gene expressed from the inactive X chromosome.

Abstract: IN mammals, equal dosage of gene products encoded by the X chromosome in male and female cells is achieved by X inactivation. Although X-chromosome inactivation represents the most extensive example known of long range cis gene regulation, the mechanism by which thousands of genes on only one of a pair of identical chromosomes are turned off is poorly understood. We have recently identified a human gene (XIST) exclusively expressed from the inactive X chromosome1. Here we report the isolation and characterization of its murine homologue (Xist) which localizes to the mouse X inactivation centre region and is the first murine gene found to be expressed from the inactive X chromosome. Nucleotide sequence analysis indicates that Xist may be associated with a protein product. The similar map positions and expression patterns for Xist in mouse and man suggest that this gene may have a role in X inactivation.

Improved molecular diagnostics for ornithine transcarbamylase deficiency.

Abstract: Since the cloning of the cDNA for X-linked ornithine transcarbamylase (OTC) in 1984, diagnostic accuracy of OTC deficiency for prenatal and carrier detection has been greatly improved by the use of linkage analysis. However, the use of RFLP-based diagnosis is limited in this and in other new mutation diseases. Here we report both the use of direct mutation detection by new PCR-based techniques and our experience with linkage-based diagnosis in 18 families. We have previously reported the use of chemical mismatch cleavage to detect mutations first in amplified mRNA and then in genomic DNA of patients. This technique has now been utilized for prenatal diagnosis. Primers for specific amplification of OTC exons 1, 3, 5, 9, and 10 have been developed and been employed to map deletions of the OTC gene in two families. These primers also have been used to detect alterations in the TaqI sites found in exons 1, 3, 5, and 9. Four novel mutations of the OTC gene leading to abolition of a TaqI site in the OTC cDNA were discovered. One of these mutations is in exon 1; two lie in exon 3; and one is in exon 9. In addition, we have used the PCR products as probes to identify the exon-specific bands seen on Southern blots and to map the polymorphic BamHI and MspI sites, which are commonly used for linkage analysis. This information will facilitate the interpretation of altered band patterns seen in deletion cases and in cases of point mutations affecting restriction sites. Utilization of the appropriate combination of these molecular techniques permitted accurate diagnostic evaluations in 17 of 18 families.

Fidelity of targeted recombination in human fibroblasts and murine embryonic stem cells

Abstract: Targeted recombination in murine embryonic stem cells promises to be a powerful tool for introducing specific mutations into target genes to study development in mice and to create animal models of human disease Gene targeting also holds potential for correcting genetic defects as an approach to human gene therapy To precisely modify target genes, homologous recombination must proceed with high fidelity However, several results have suggested that targeted recombination may be highly mutagenic To test the accuracy of gene targeting we analyzed 44 independent targeted recombinants at the hypoxanthine phosphoribosyltransferase (HPRT) locus in a human fibroblast cell line and in mouse embryonic stem cells We surveyed 80 kilobases around the sites of recombination by using chemical cleavage of mismatches Only two mutations were found: a T----G transversion and a thymidine deletion Thus, gene targeting in mammalian cells can be extremely accurate These results demonstrate the feasibility of generating precise modifications of mammalian genomes by gene targeting

Gene therapy in man and mice: adenosine deaminase deficiency, ornithine transcarbamylase deficiency, and Duchenne muscular dystrophy.

Abstract: Gene therapy is defined as the delivery and expression of a functional gene into somatic tissues of patients (or animals) endogenously deficient in this gene. Depending on the natural tissue pattern of expression and the sites most affected by disease processes, different anatomic sites may be the targets for gene transduction. To date, retrovirus-mediated gene transfer into hemopoietic stem cells is the most advanced of gene therapy systems and the most promising in terms of clinical applications in the near future. There are, however, additional efforts under way in our laboratory to develop efficient means of transducing genes into the liver, intestine, skeletal and cardiac muscles.