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Soria L.A.,University of Buenos Aires | Corva P.M.,University of the Sea | Huguet M.J.,University of Buenos Aires | Mino S.,Institute Virologia | Miquel M.C.,University of Buenos Aires
BAG - Journal of Basic and Applied Genetics

The bovine CAPN1 gene encodes the large subunit of μ-calpain, which is thought to be one of the most important enzymes involved in postmortem beef tenderization. Three SNPs in CAPN1 (SNP 316, SNP 530 and SNP 4751) have been associated with beef tenderness in different beef cattle breeds. The objective of this work was to implement genotyping strategies for CAPN1 markers as part of a project pursuing the identifi cation and validation of molecular markers associated with bovine meat quality and composition. Three PCR-RFLP methods were designed to determine genotypes of 64 bulls (11 Angus, 43 Brangus and 10 Brahman). Unexpected patterns resulting from the PCR-RFLP analysis at SNP 316 and SNP 530 were resolved by cloning and sequencing and lead to the discovery of three substitutions not previously described. These mutations could be useful in population studies, such as the determination of the relative contribution of the Angus and Brahman breeds to the Brangus. Source

Pauly D.,Robert Koch Institute | Chacana P.A.,Institute Virologia | Brembs B.,Free University of Berlin | Schade R.,Charite - Medical University of Berlin
Journal of Visualized Experiments

Hens can be immunized by means of i.m. vaccination (Musculus pectoralis, left and right, injection volume 0.5-1.0 ml) or by means of Gene-Gun plasmid-immunization. Dependent on the immunogenicity of the antigen, high antibody-titres (up to 1:100,000 - 1:1,000,000) can be achieved after only one or 3 - 4 boost immunizations. Normally, a hen lays eggs continuously for about 72 weeks, thereafter the laying capacity decreases. This protocol describes the extraction of total IgY from egg yolk by means of a precipitation procedure (PEG. Polson et al. 1980). The method involves two important steps. The first one is the removal of lipids and the second is the precipitation of total IgY from the supernatant of step one. After dialysis against a buffer (normally PBS) the IgY-extract can be stored at -20°C for more than a year. The purity of the extract is around 80%, the total IgY per egg varies from 40-80 mg, dependent on the age of the laying hen. The total IgY content increases with the age of the hen from around 40 mg/egg up to 80 mg/egg (concerning PEG precipitation). The laying capacity of a hen per year is around 325 eggs. That means a total potential harvest of 20 g total IgY/year based on a mean IgY content of 60 mg total IgY/egg (see Table 1). © 2011 Journal of Visualized Experiments. Source

Gillet N.A.,University of Liege | Hamaidia M.,University of Liege | de Brogniez A.,University of Liege | Gutierrez G.,Institute Virologia | And 4 more authors.
PLoS Pathogens

Retroviruses are not expected to encode miRNAs because of the potential problem of self-cleavage of their genomic RNAs. This assumption has recently been challenged by experiments showing that bovine leukemia virus (BLV) encodes miRNAs from intragenomic Pol III promoters. The BLV miRNAs are abundantly expressed in B-cell tumors in the absence of significant levels of genomic and subgenomic viral RNAs. Using deep RNA sequencing and functional reporter assays, we show that miRNAs mediate the expression of genes involved in cell signaling, cancer and immunity. We further demonstrate that BLV miRNAs are essential to induce B-cell tumors in an experimental model and to promote efficient viral replication in the natural host. © 2016 Gillet et al. Source

Legisa D.M.,Institute Virologia | Perez Aguirreburualde M.S.,Institute Virologia | Gonzalez F.N.,Institute Virologia | Marin-Lopez A.,Research Center en Sanidad Animal | And 4 more authors.

Bluetongue virus (BTV), the causative agent of bluetongue disease (BT) in domestic and wild ruminants, is worldwide distributed. A total of 27 serotypes have been described so far, and several outbreaks have been reported. Vaccination is critical for controlling the spread of BTV. In the last years, subunit vaccines, viral vector vaccines and reverse genetic-based vaccines have emerged as new alternatives to conventional ones. In this study, we developed an experimental subunit vaccine against BTV4, with the benefit of targeting the recombinant protein to antigen-presenting cells. The VP2 protein from an Argentine BTV4 isolate was expressed alone or fused to the antigen presenting cell homing (APCH) molecule, in the baculovirus insect cell expression system. The immunogenicity of both proteins was evaluated in guinea pigs and cattle. Titers of specific neutralizing antibodies in guinea pigs and cattle immunized with VP2 or APCH-VP2 were high and similar to those induced by a conventional inactivated vaccine. The immunogenicity of recombinant proteins was further studied in the IFNAR(-/-) mouse model where the fusion of VP2 to APCH enhanced the cellular immune response and the neutralizing activity induced by VP2. © 2015 Elsevier Ltd. Source

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