ARTS: a web-based tool for the set-up of high-throughput genome-wide mapping panels for the SNP genotyping of mouse mutants

Genome-wide mapping in the identification of novel candidate genes has always been the standard method in genetics and genomics to correlate a clinically interesting phenotypic trait with a genotype. However, the performance of a mapping experiment using classical microsatellite approaches can be very time consuming. The high-throughput analysis of single-nucleotide polymorphisms (SNPs) has the potential of being the successor of microsatellite analysis routinely used for these mapping approaches, where one of the major obstacles is the design of the appropriate SNP marker set itself. Here we report on ARTS, an advanced retrieval tool for SNPs, which allows researchers to comb freely the public mouse dbSNP database for multiple reference and test strains. Several filters can be applied in order to improve the sensitivity and the specificity of the search results. By employing the panel generator function of this program, it is possible to abbreviate the extraction of reliable sequence data for a large marker panel including several different mouse strains from days to minutes. The concept of ARTS is easily adaptable to other species for which SNP databases are available, making it a versatile tool for the use of SNPs as markers for genotyping. The web interface is accessible at

A novel concept for the genome wide high-throughput mapping of ENU-induced mouse mutants

Ziel der Arbeit war die Erstellung einer genomweiten SNP-Genotypisierungsplattform für die Kartierung von ENU-induzierten Mausmutanten. Im Zuge dessen wurde eine Reihe von bioinformatischen Werkzeugen für die Erstellung eines geeigneten Markersatzes und für die Auswertung der gewonnenen Genotypdaten entwickelt. Mit Hilfe dieser Plattform war es möglich, eine genomweite Kartierung von 55 mutanten Mauslinien durchzuführen. Der durchschnittliche Abstand des von der ENU Mutagenese betroffenen Gens zu dem am stärksten gekoppelten SNP-Marker betrug hierbei ca. 5,9 Mb. Eine der auf diese Weise untersuchten mutanten Mauslinien war die Linie SMA004. Es wurde eine Kopplung auf Chromosom 4 festgestellt und eine Mutation in dem Gen Zmpste24 gefunden. Der progeroide Phänotyp dieser Mauslinie wurde weiterführend untersucht. In dieser Arbeit wird eine neuartige Plattform präsentiert, mit deren Hilfe es möglich ist, in kurzer Zeit Kreuzungen verschiedener Mausstämme zu genotypisieren und auszuwerten.Aim of this work was the establishment of a genome wide SNP genotyping platform for the mapping of ENU-induced mouse mutants. During this course a set of bioinformatic tools for the establishment of a suitable marker set and for the analysis of the obtained genotype data has been developed. By means of this platform it was possible to perform a genome wide mapping of 55 mutant mouse lines. The average distance between the gene alted by ENU mutagenesis and the most strongly linked SNP-marker was thereby approximately 5,9 Mb. One of the mouse lines assayed in this manner was the line SMA004. A linkage on chromosome 4 was observed and a mutation in the gene Zmpste24 has been found. The progeroid phenotype of this mouse line was further studied. In this thesis a novel platform is presented by means of which it is possible to genotype and analyze crosses of different mouse strains within a short time

Dominant-negative effects of a novel mutated Ins2 allele causes early-onset diabetes and severe beta-cell loss in Munich Ins2C95S mutant mice

The novel diabetic mouse model Munich Ins2(C95S) was discovered within the Munich N-ethyl-N-nitrosourea mouse mutagenesis screen. These mice exhibit a T-->A transversion in the insulin 2 (Ins2) gene at nucleotide position 1903 in exon 3, which leads to the amino acid exchange C95S and loss of the A6-A11 intrachain disulfide bond. From 1 month of age onwards, blood glucose levels of heterozygous Munich Ins2(C95S) mutant mice were significantly increased compared with controls. The fasted and postprandial serum insulin levels of the heterozygous mutants were indistinguishable from those of wild-type littermates. However, serum insulin levels after glucose challenge, pancreatic insulin content, and homeostasis model assessment (HOMA) beta-cell indices of heterozygous mutants were significantly lower than those of wild-type littermates. The initial blood glucose decrease during an insulin tolerance test was lower and HOMA insulin resistance indices were significantly higher in mutant mice, indicating the development of insulin resistance in mutant mice. The total islet volume, the volume density of beta-cells in the islets, and the total beta-cell volume of heterozygous male mutants was significantly reduced compared with wild-type mice. Electron microscopy of the beta-cells of male mutants showed virtually no secretory insulin granules, the endoplasmic reticulum was severely enlarged, and mitochondria appeared swollen. Thus, Munich Ins2(C95S) mutant mice are considered a valuable model to study the mechanisms of beta-cell dysfunction and death during the development of diabetes

A Genetic Screen for Modifiers of the Delta1-Dependent Notch Signaling Function in the Mouse

The Notch signaling pathway is an evolutionarily conserved transduction pathway involved in embryonic patterning and regulation of cell fates during development. Recent studies have demonstrated that this pathway is integral to a complex system of interactions, which are also involved in distinct human diseases. Delta1 is one of the known ligands of the Notch receptors. Mice homozygous for a loss-of-function allele of the Delta1 gene Dll1(lacZ/lacZ) die during embryonic development. Here, we present the results of two phenotype-driven modifier screens. Heterozygous Dll1(lacZ) knockout animals were crossed with ENU-mutagenized mice and screened for dysmorphological, clinical chemical, and immunological variants that are dependent on the Delta1 loss-of-function allele. First, we show that mutagenized heterozygous Dll1(lacZ) offspring have reduced body weight and altered specific clinical chemical parameters, including changes in metabolites and electrolytes relevant for kidney function. In our mutagenesis screen we have successfully generated 35 new mutant lines. Of major interest are 7 mutant lines that exhibit a Dll1(lacZ/+)-dependent phenotype. These mutant mouse lines provide excellent in vivo tools for studying the role of Notch signaling in kidney and liver function, cholesterol and iron metabolism, cell-fate decisions, and during maturation of T cells in the immune system

New C3H Kit(N824K/WT) cancer mouse model develops late-onset malignant mammary tumors with high penetrance

Gastro-intestinal stromal tumors and acute myeloid leukemia induced by activating stem cell factor receptor tyrosine kinase (KIT) mutations are highly malignant. Less clear is the role of KIT mutations in the context of breast cancer. Treatment success of KIT-induced cancers is still unsatisfactory because of primary or secondary resistance to therapy. Mouse models offer essential platforms for studies on molecular disease mechanisms in basic cancer research. In the course of the Munich N-ethyl-N-nitrosourea (ENU) mutagenesis program a mouse line with inherited polycythemia was established. It carries a base-pair exchange in the Kit gene leading to an amino acid exchange at position 824 in the activation loop of KIT. This KIT variant corresponds to the N822K mutation found in human cancers, which is associated with imatinib-resistance. C3H Kit(N824K/WT) mice develop hyperplasia of interstitial cells of Cajal and retention of ingesta in the cecum. In contrast to previous Kit-mutant models, we observe a benign course of gastrointestinal pathology associated with prolonged survival. Female mutants develop mammary carcinomas at late onset and subsequent lung metastasis. The disease model complements existing oncology research platforms. It allows for addressing the role of KIT mutations in breast cancer and identifying genetic and environmental modifiers of disease progression

Immuno-histochemistry ERCC2 expression.

<p>Using antibodies against ERCC2 and Alexa488 (green) as secondary antibody, we demonstrate that ERCC2 is expressed weakly in the ocular lens around birth; expression is increasing from P7 onward. Cell nuclei are counterstained by DAPI (blue). ERCC2 is mainly expressed in the cytosol.</p

Primers for PCR.

<p>Primers for PCR.</p

<i>RCO015</i> mutants and identification of the underlying mutation.

<p>a) A heterozygous (left) and homozygous (middle) <i>RCO015</i> mutant mouse at the age of 9 months compared to a wild type (right). The homozygous mutants are smaller and have rough hair and small eyes. b) Haplotype analysis defines the critical interval between the markers <i>116J6</i>.<i>1</i> and <i>D7Mit294</i> at mouse chromosome 7.</p