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Aeromonas hydrophila is a major bacterial pathogen associated with hemorrhagic septicemia in aquatic and terrestrial animals including humans. There is an urgent need to develop molecular and immunological assays for rapid, specific and sensitive diagnosis. A new set of primers has been designed for detection of thermostable hemolysin (TH) gene (645 bp) from A. hydrophila, and sensitivity limit for detection of TH gene was 5 pg. The TH gene was cloned, sequenced and analyzed. The G+C content was 68.06%; and phylogeny was constructed using TH protein sequences which had significant homology with those for thermostable and other hemolysins present in several bacterial pathogens. In addition, we have predicted the four and eight T-cell epitopes for MHC class I and II alleles, respectively. These results provide new insight for TH protein containing antigenic epitopes that can be used in immunoassays and also designing of thermostable vaccines. Source

The engineering of synthetic gene networks has mostly relied on the assembly of few characterized regulatory elements using rational design principles. It is of outmost importance to analyze the scalability and limits of such a design workflow. To analyze the design capabilities of libraries of regulatory elements, we have developed the first automated design approach that combines such elements to search the genotype space associated to a given phenotypic behavior. Herein, we calculated the designability of dynamical functions obtained from circuits assembled with a given genetic library. By designing circuits working as amplitude filters, pulse counters and oscillators, we could infer new mechanisms for such behaviors. We also highlighted the hierarchical design and the optimization of the interface between devices. We dissected the functional diversity of a constrained library and we found that even such libraries can provide a rich variety of behaviors. We also found that intrinsic noise slightly reduces the designability of digital circuits, but it increases the designability of oscillators. Finally, we analyzed the robust design as a strategy to counteract the evolvability and noise in gene expression of the engineered circuits within a cellular background, obtaining mechanisms for robustness through non-linear negative feedback loops. Source

Jordan B.R.,Genopole
Expert Review of Molecular Diagnostics | Year: 2010

DNA microarrays, 15 years after their appearance, have achieved presence in a number of medical settings. Several tests have been introduced and have obtained regulatory approval, mostly in the fields of bacterial identification, mutation detection and the global assessment of genome alterations, a particularly successful case being the whole-genome assay of copy-number variations. Gene-expression applications have been less successful because of technical issues (e.g., reproducibility, platform-to-platform consistency and statistical issues in data analysis) and difficulties in demonstrating the clinical utility of expression signatures. In their different applications, DNA arrays have faced competition from PCR-based assays for low and intermediate multiplicity. Now they have a new competitor, new-generation sequencing, that can provide a wealth of direct sequence information, or digital gene-expression data, at a constantly decreasing cost. In this article we evaluate the strengths and weaknesses of the DNA microarray approach to diagnostics, and highlight the fields in which it is most likely to achieve a durable presence. © 2010 Expert Reviews Ltd. Source

Tempel S.,Genopole
Methods in Molecular Biology | Year: 2012

RepeatMasker is a program that screens DNA sequences for interspersed repeats and low-complexity DNA sequences. In this chapter, we present the procedure to routinely use this program on a personal computer. © 2012 Springer Science+Business Media, LLC. Source

Tempel S.,Genopole | Tahi F.,Genopole
Nucleic Acids Research | Year: 2012

miRNAs are small non coding RNA structures which play important roles in biological processes. Finding miRNA precursors in genomes is therefore an important task, where computational methods are required. The goal of these methods is to select potential pre-miRNAs which could be validated by experimental methods. With the new generation of sequencing techniques, it is important to have fast algorithms that are able to treat whole genomes in acceptable times. We developed an algorithm based on an original method where an approximation of miRNA hairpins are first searched, before reconstituting the pre-miRNA structure. The approximation step allows a substantial decrease in the number of possibilities and thus the time required for searching. Our method was tested on different genomic sequences, and was compared with CID-miRNA, miRPara and VMir. It gives in almost all cases better sensitivity and selectivity. It is faster than CID-miRNA, miRPara and VMir: it takes ∼30s to process a 1 MB sequence, when VMir takes 30min, miRPara takes 20h and CID-miRNA takes 55h. We present here a fast ab-initio algorithm for searching for pre-miRNA precursors in genomes, called miRNAFold. miRNAFold is available at http://EvryRNA.ibisc.univ-evry.fr/. © 2012 The Author(s). Source

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