High Resolution Melt Analysis (HRMA); a Viable Alternative to Agarose Gel Electrophoresis for Mouse Genotyping

Most mouse genetics laboratories maintain mouse strains that require genotyping in order to identify the genetically modified animals. The plethora of mutagenesis strategies and publicly available mouse alleles means that any one laboratory may maintain alleles with random or targeted insertions of orthologous or unrelated sequences as well as random or targeted deletions and point mutants. Many experiments require that different strains be cross bred conferring the need to genotype progeny at more than one locus. In contrast to the range of new technologies for mouse mutagenesis, genotyping methods have remained relatively static with alleles typically discriminated by agarose gel electrophoresis of PCR products. This requires a large amount of researcher time. Additionally it is susceptible to contamination of future genotyping experiments because it requires that tubes containing PCR products be opened for analysis. Progress has been made with the genotyping of mouse point mutants because a range of new high-throughput techniques have been developed for the detection of Single Nucleotide Polymorphisms. Some of these techniques are suitable for genotyping point mutants but do not detect insertion or deletion alleles. Ideally, mouse genetics laboratories would use a single, high-throughput platform that enables closed-tube analysis to genotype the entire range of possible insertion and deletion alleles and point mutants. Here we show that High Resolution Melt Analysis meets these criteria, it is suitable for closed-tube genotyping of all allele types and current genotyping assays can be converted to this technology with little or no effort.This work was supported by NHMRC (http://www.nhmrc.gov.au) grants 366476 and 1003648 (to R.M.A.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Energy deposition simulations for a damage experiment with superconducting sample coils

An experiment to study damage caused by the impact of 440 GeV/c protons on sample superconducting racetrack coils made from NbTi and Nb3Sn strands was recently carried out at CERN\u27s HiRadMat facility. This paper reports on the detailed Monte Carlo simulations performed with FLUKA and Geant4 to evaluate the energy deposition of the 440 GeV/c proton beam on the sample coils positioned in the experimental setup. using the measured beam parameters during the experiment. The measured hotspot temperatures and temperature gradients reached in the sample coils are presented and compared with the simulations. In addition, comparisons between the simulation results from FLUKA and Geant4 are discussed in detail

Immunoglobulin G structure and rheumatoid factor epitopes

Antibodies are important for immunity and exist in several classes (IgM, IgD, IgA, IgG, IgE). They are composed of symmetric dimeric molecules with two antigen binding regions (Fab) and a constant part (Fc), usually depicted as Y-shaped molecules. Rheumatoid factors found in patients with rheumatoid arthritis are autoantibodies binding to IgG and paradoxically appear to circulate in blood alongside with their antigen (IgG) without reacting with it. Here, it is shown that rheumatoid factors do not react with native IgG in solution, and that their epitopes only become accessible upon certain physico-chemical treatments (e.g. heat treatment at 57 °C), by physical adsorption on a hydrophobic surface or by antigen binding. Moreover, chemical cross-linking in combination with mass spectrometry showed that the native state of IgG is a compact (closed) form and that the Fab parts of IgG shield the Fc region and thereby control access of rheumatoid factors and presumably also some effector functions. It can be inferred that antibody binding to pathogen surfaces induces a conformational change, which exposes the Fc part with its effector sites and rheumatoid factor epitopes. This has strong implications for understanding antibody structure and physiology and necessitates a conceptual reformulation of IgG models

Dramaturger om dramaturgi: enquete

Ten Danish dramatrugs from different theatrical fields tell about their daily work and experoences, and their visions for the changing role of the dramaturg in contemporary theatr

Host Immune Transcriptional Profiles Reflect the Variability in Clinical Disease Manifestations in Patients with Staphylococcus aureus Infections

Staphylococcus aureus infections are associated with diverse clinical manifestations leading to significant morbidity and mortality. To define the role of the host response in the clinical manifestations of the disease, we characterized whole blood transcriptional profiles of children hospitalized with community-acquired S. aureus infection and phenotyped the bacterial strains isolated. The overall transcriptional response to S. aureus infection was characterized by over-expression of innate immunity and hematopoiesis related genes and under-expression of genes related to adaptive immunity. We assessed individual profiles using modular fingerprints combined with the molecular distance to health (MDTH), a numerical score of transcriptional perturbation as compared to healthy controls. We observed significant heterogeneity in the host signatures and MDTH, as they were influenced by the type of clinical presentation, the extent of bacterial dissemination, and time of blood sampling in the course of the infection, but not by the bacterial isolate. System analysis approaches provide a new understanding of disease pathogenesis and the relation/interaction between host response and clinical disease manifestations