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 phylogenetic framework for the North American bumblebee species of the subgenus Bombus sensu stricto (Bombus affinis, B. franklini, B. moderatus, B. occidentalis & B. terricola) based on mitochondrial DNA markers. (Hymenoptera: Apidae: Bombus).

Königinnen der vier Taxa Bombus affinis, B. moderatus, B. occidentalis and B. terricola wurden an verschiedenen Orten quer durch Nordamerika gefangen. Zusätzlich wurden Männchen von B. franklini, B. occidentalis und B. terricola gesammelt. Mitochondriale Cytochrome Oxidase Untereinheit I (COI) von 25 Proben wurde sequenziert (Teilsequenzen 1005 bp Länge). Die Divergenz der Sequenzen zwischen den Taxa beträgt 30–50 Basen-Substitutionen und die Tamura-Nei Genetische Distanz 0.05–0.13, während innerhalb der Taxa die Divergenz nur 1 bis 2 Basen-Substitutionen beträgt und die Tamura-Nei Genetische Distanz 0.001–0.002. Da die COI Sequenzen keine Lücken aufweisen, können die einzelnen Nukleotide wie homologe Positionen verwendet werden. Jedes Taxon besitzt 8–20 eigene Substitutionen, die als dia­gnostische Positionen verwendet werden können, um das Taxon zu charakterisieren. Das Phylogramm zeigt drei klar getrennte Cluster: B. moderatus, konspezifisch mit der ostasiatischen B. albocinctus, das Artenpaar B. affinis – B. franklini locker verwandt mit der eurasiatischen B. lucorum, und das Artenpaar B. terricola – B. occidentalis ohne Beziehungen zu irgendwelchen Arten der Alten Welt.Queens of Bombus affinis, B. moderatus, B. occidentalis and B. terricola were collected from different localities throughout North America. In addition, males of B. franklini, B. occidentalis and B. terricola were collected. Mitochondrial cytochrome oxidase subunit I (COI) was sequenced from 24 specimens (Partial sequences length 1005 bp). Interspecific sequence divergence was about 30–50 base substitutions and approximately 0.05–0.13 in Tamura-Nei genetic distance, whereas the intraspecific sequence divergence was only 1–2 base substitutions and about 0.001–0.002 in Tamura-Nei genetic distance. Because there are no gaps in the alignments of the COI sequences, single nucleotide sites can be used as positional homologies. Each taxon is characterised by about 8 to 20 substitutions, which are unique (“private”) and can be used as diagnostic characters to define and identify this taxon. Three clusters in the topology of the phylogenetic tree were obtained: B. moderatus, which is con-specific to the East Asian B. albocinctus, a species pair B. affinis – B. franklini somehow related to the Eurasian B. lucorum and a species pair B. terricola – B. occidentalis without obvious connection to any Old World species

Haplotypes, median networks, and diagnostic characters as tools to elucidate the intraspecific genetic and taxonomic structure of bumblebees, with a description of Bombus cryptarum pallidocinctus new subspecies (Hymenoptera: Apidae: Bombus).

Ein Alignment von 168 Sequenzen mitochondrialer DNA von Bombus cryptarum liefert 29 Haplotypen. Ein Median-Network zeigt fünf klar abgrenzbare Cluster, die Haplogruppen (HG) A-E, die durch diagnostische Positionen definiert werden. Vier dieser HG lassen sich bekannten Taxa zuordnen HG-A = B. cryptarum cryptarum, HG-B = B. cryptarum florilegus, HG-D = B. cryptarum albocinctus, und HG-E = B. cryptarum moderatus. Die Haplogruppe C entspricht einem neuen Taxon B. cryptarum pallidocinctus im Rang einer Unterart. Diese neue Unterart wird beschrieben. Die Verbreitung der Eurasiatischen Taxa wird untersucht und die Folgerungen aus Median-Network und geographischer Verbreitung für die mögliche Phylogenie werden diskutiert.StichwörterHymenoptera, Apidae, Bombus, mitochondrial DNA, new subspecies, geographic distributionNomenklatorische Handlungencryptarum pallidocinctus Bertsch et al., 2014 (Bombus), subspec. n.An alignment of 168 sequences of mitochondrial DNA of Bombus cryptarum reveals 29 haplotypes. A median network shows five distinct clusters, haplogroups (HG) A-E, which can be defined by diagnostic characters. Four of these HGs can be assigned to well-known taxa: HG-A = B. cryptarum cryptarum; HG-B = B. cryptarum florilegus; HG-D = B. cryptarum albocinctus; and HG-E = B. cryptarum moderatus. Haplogroup C represents a new taxon B. cryptarum pallidocinctus in the rank of a subspecies. This new subspecies is described. The geographic distribution of the Eurasiatic taxa is described and the implications of geographic distribution and median network for the phylogeny are discussed.KeywordsHymenoptera, Apidae, Bombus, mitochondrial DNA, new subspecies, geographic distributionNomenclatural Actscryptarum pallidocinctus Bertsch et al., 2014 (Bombus), subspec. n

Phylogenetic relationships of the bumblebees Bombus moderatus, B. albocinctus, B. burjaeticus, B. florilegus and B. cryptarum based on mitochondrial DNA markers: a complex of closely related taxa with circumpolar distribution. (Hymenoptera: Apidae: Bombus).

Königinnen von Bombus moderatus aus Alaska/USA und Alberta/Canada und von B. burjaeticus and B. patagiatus aus dem Russischen Transbaikal wurden an verschiedenen Orten im Frühjahr gefangen, um künstliche Kolonien zu züchten. Zusätzlich wurden Männchen von B. florilegus in Hokkaido/Japan gesammelt. Teilsequenzen (Länge 1005 bp) mitochondrialer Cytochrome Oxidase Untereinheit I (COI) wurde sequenziert. Zum Vergleich wurden auch Museumsproben von B. albocinctus und B. burjaeticus sequenziert. Die Divergenz der Sequenzen innerhalb der Taxa beträgt 1 bis 2 Basen-Substitutionen und die Tamura-Nei Genetische Distanz 0.001–0.002. Die Divergenz der Sequenzen zwischen B. moderatus, B. albocinctus und B. burjaeticus beträgt nur 1–5 Basen-Substitutionen und die Tamura-Nei Genetische Distanz 0.001–0.005, während die Divergenz der Sequenzen zwischen B. lucorum, B. magnus, B. patagiatus und B. cryptarum 22-44 Basen Substitutionen beträgt und die Tamura-Nei Genetische Distanz 0.027-0.042. Zusätzlich zu den Clustern für B. lucorum, B. magnus und B. patagiatus zeigt die Topologie des Phylogramms (MrBayes Maximum Likelihood Tree) ein umfangreiches Cluster, in dem B. albocinctus, B. burjaeticus, B. moderatus und B. florilegus vereinigt sind. Da die COI Sequenzen keine Lücken aufweisen, können die einzelnen Nukleotide wie homologe Positionen verwendet werden. Jedes Taxon besitzt 8–20 eigene Substitutionen, die als diagnostische Positionen zur Charakterisierung des Taxons verwendet werden können. Die Analyse der diagnostischen Positionen der Taxa bestätigt die Topologie des Maximum Likelihood Phylogramms. Um die Lücke zwischen den östlichsten bekannten Vorkommen von B. cryptarum im Kaukasus und im Elburz und den Vorkommen von B. burjaeticus/ B. albocinctus im Russischen Transbaikal und in Russisch Fernost zu überbrücken, wurden 12 weitere Museumsproben aus Zentralasiatischen Gebirgen und dem Himalaja sequenziert. Durch Analyse der diagnostischen Positionen der teilweise 100 Jahre alten DNA kann gezeigt werden, dass keine dieser Proben zu B. lucorum gehören kann, alle bilden ein Cluster mit den Taxa des cryptarum Komplexes.Spring queens of Bombus moderatus from Alaska/USA and Alberta/Canada, and of B. burjaeticus and B. patagiatus from the Russian Transbaikal region were collected at different localities. In addition, males of B. florilegus were collected from Hokkaido/Japan. Partial sequences (length 1005 bp) of mitochondrial cytochrome oxidase subunit I (COI) were sequenced from specimens from each locality and species. For comparison, museum specimens of B. albocinctus and B. burjaeticus were also sequenced. The intraspecific sequence divergence was only 1–2 base substitutions and about 0.001–0.002 in Tamura-Nei genetic distance. The interspecific sequence divergence between B. moderatus, B. albocinctus and B. burjaeticus was only 1–5 base substitutions and about 0.001–0.005 in Tamura-Nei genetic distance, whereas the sequence divergence between B. lucorum, B. magnus, B. patagiatus and B. cryptarum was about 24–44 base substitutions and approximately 0.027–0.042 in Tamura-Nei genetic distance. The MrBayes maximum likelihood tree generated a tree topology with three separate clusters for B. lucorum, B. magnus and B. patagiatus, and one large cluster which united B. albocinctus, B. burjaeticus, B. moderatus and B. florilegus. Because there are no gaps in the alignments of COI sequences, single nucleotide sites were used as positional homologies. Each taxon was characterised by about 8–20 substitutions which were unique (“private”) and could be used as diagnostic characters to define and identify these taxa. An analysis of the number of diagnostic characters confirmed the clustering of the maximum likelihood tree. To bridge the gap of about 5000 km between the most eastern known localities for B. cryptarum in the Caucasus and Elburz Mountains and B. burjaeticus/ B. albocinctus in the Russian Transbaikal region and the Russian Far East, 12 more museum specimens from the Central Asiatic Mountains and the Himalayas were sequenced. By analysing the diagnostic positions in the sequences of this almost 100-year-old museum DNA, it was shown that none were connected with B. lucorum: they all clustered within the cryptarum-complex taxa

Compartmentalised expression of Delta-like 1 in epithelial somites is required for the formation of intervertebral joints

<p>Abstract</p> <p>Background</p> <p>Expression of the mouse <it>Delta-like 1 </it>(<it>Dll1</it>) gene in the presomitic mesoderm and in the caudal halves of somites of the developing embryo is required for the formation of epithelial somites and for the maintenance of caudal somite identity, respectively. The rostro-caudal polarity of somites is initiated early on within the presomitic mesoderm in nascent somites. Here we have investigated the requirement of restricted <it>Dll1 </it>expression in caudal somite compartments for the maintenance of rostro-caudal somite polarity and the morphogenesis of the axial skeleton. We did this by overexpressing a functional copy of the <it>Dll1 </it>gene throughout the paraxial mesoderm, in particular in anterior somite compartments, during somitogenesis in transgenic mice.</p> <p>Results</p> <p>Epithelial somites were generated normally and appeared histologically normal in embryos of two independent <it>Dll1 </it>over-expressing transgenic lines. Gene expression analyses of rostro-caudal marker genes suggested that over-expression of <it>Dll1 </it>without restriction to caudal compartments was not sufficient to confer caudal identity to rostral somite halves in transgenic embryos. Nevertheless, <it>Dll1 </it>over-expression caused dysmorphologies of the axial skeleton, in particular, in morphological structures that derive from the articular joint forming compartment of vertebrae. Accordingly, transgenic animals exhibited missing or reduced intervertebral discs, rostral and caudal articular processes as well as costal heads of ribs. In addition, the midline of the vertebral column did not develop normally. Transgenic mice had open neural arches and split vertebral bodies with ectopic pseudo-growth plates. Endochondral bone formation and ossification in the developing vertebrae were delayed.</p> <p>Conclusion</p> <p>The mice overexpressing <it>Dll1 </it>exhibit skeletal dysmorphologies that are also evident in several mutant mice with defects in somite compartmentalisation. The <it>Dll1 </it>transgenic mice demonstrate that vertebral dysmorphologies such as bony fusions of vertebrae and midline vertebral defects can occur without apparent changes in somitic rostro-caudal marker gene expression. Also, we demonstrate that the over-expression of the <it>Dll1 </it>gene in rostral epithelial somites is not sufficient to confer caudal identity to rostral compartments. Our data suggest that the restricted <it>Dll1 </it>expression in caudal epithelial somites may be particularly required for the proper development of the intervertebral joint forming compartment.</p

Linking disease-associated genes to regulatory networks via promoter organization

Pathway- or disease-associated genes may participate in more than one transcriptional co-regulation network. Such gene groups can be readily obtained by literature analysis or by high-throughput techniques such as microarrays or protein-interaction mapping. We developed a strategy that defines regulatory networks by in silico promoter analysis, finding potentially co-regulated subgroups without a priori knowledge. Pairs of transcription factor binding sites conserved in orthologous genes (vertically) as well as in promoter sequences of co-regulated genes (horizontally) were used as seeds for the development of promoter models representing potential co-regulation. This approach was applied to a Maturity Onset Diabetes of the Young (MODY)-associated gene list, which yielded two models connecting functionally interacting genes within MODY-related insulin/glucose signaling pathways. Additional genes functionally connected to our initial gene list were identified by database searches with these promoter models. Thus, data-driven in silico promoter analysis allowed integrating molecular mechanisms with biological functions of the cell

Carbon-nanoparticle-triggered acute lung inflammation and its resolution are not altered in PPARγ-defective (P465L) mice.

BACKGROUND: The alveolar macrophage (AM) - first line of innate immune defence against pathogens and environmental irritants - constitutively expresses peroxisome-proliferator activated receptor γ (PPARγ). PPARγ ligand-induced activation keeps the AM quiescent, and thereby contributes to combat invaders and resolve inflammation by augmenting the phagocytosis of apoptotic neutrophils and inhibiting an excessive expression of inflammatory genes. Because of these presumed anti-inflammatory functions of PPARγ we tested the hypothesis, whether reduced functional receptor availability in mutant mice resulted in increased cellular and molecular inflammatory response during acute inflammation and/or in an impairment of its resolution. METHODS: To address this hypothesis we examined the effects of a carbon-nanoparticle (CNP) lung challenge, as surrogate for non-infectious environmental irritants, in a murine model carrying a dominant-negative point mutation in the ligand-binding domain of PPARγ (P465L/wt). Animals were instilled intratracheally with Printex 90 CNPs and bronchoalveolar lavage (BAL) was gained 24 h or 72 h after instillation to investigate its cellular and protein composition. RESULTS: Higher BAL cell numbers - due to higher macrophage counts - were found in mutants irrespective of treatment. Neutrophil numbers in contrast were slightly lower in mutants. Intratracheal CNP instillation resulted in a profound recruitment of inflammatory neutrophils into the alveolus, but genotype related differences at acute inflammation (24 h) and resolution (72 h) were not observed. There were no signs for increased alveolar-capillary membrane damage or necrotic cell death in mutants as determined by BAL protein and lactate-dehydrogenase content. Pro-inflammatory macrophage-derived cytokine osteopontin was higher, but galectin-3 lower in female mutants. CXCL5 and lipocalin-2 markers, attributed to epithelial cell stimulation did not differ. CONCLUSIONS: Despite general genotype-related differences, we had to reject our hypothesis of an increased CNP induced lung inflammation and an impairment of its resolution in PPARγ defective mice. Although earlier studies showed ligand-induced activation of nuclear receptor PPARγ to promote resolution of lung inflammation, its reduced activity did not provide signs of resolution impairment in the settings investigated here.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

MausDB: An open source application for phenotype data and mouse colony management in large-scale mouse phenotyping projects

<p>Abstract</p> <p>Background</p> <p>Large-scale, comprehensive and standardized high-throughput mouse phenotyping has been established as a tool of functional genome research by the German Mouse Clinic and others. In all these projects, vast amounts of data are continuously generated and need to be stored, prepared for data-mining procedures and eventually be made publicly available. Thus, central storage and integrated management of mouse phenotype data, genotype data, metadata and linked external data are highly important. Requirements most probably depend on the individual mouse housing unit or project and the demand for either very specific individual database solutions or very flexible solutions that can be easily adapted to local demands. Not every group has the resources and/or the know-how to develop software for this purpose. A database application has been developed for the German Mouse Clinic in order to meet all requirements mentioned above.</p> <p>Results</p> <p>We present MausDB, the German Mouse Clinic web-based database application that integrates standard mouse colony management, phenotyping workflow scheduling features and mouse phenotyping result data management. It links mouse phenotype data with genotype data, metadata and external data such as public web databases, which is a prerequisite for comprehensive data analysis and mining. We describe how this can be achieved with a lean and user-friendly system built on open standards.</p> <p>Conclusion</p> <p>MausDB is suited for large-scale, high-throughput phenotyping facilities but can also be used exclusively for mouse colony management within smaller units or projects. The system is successfully used as the primary mouse and data management tool of the German Mouse Clinic and other mouse facilities. We offer MausDB to the scientific community as open source software to provide a system for storage of data from functional genomics projects in a well-structured, easily accessible form.</p