ERROR AND UNCERTAINTY ANALYSIS OF RESIDUAL STRESS EVALUATION BY USINGTHE RING-CORE METHOD

The Ring-Core Method is a technique used for the experimental analysis of the residual stresses in mechanical components. For uniform and non-uniform residual stresses estimation, the use of the method leads in general to accurate results but, unfortunately at present the user does not have appropriate procedures to correct the obtained results from systematic errors as well as to estimate the uncertainty due to random errors. In order to overcome such drawbacks, in the present work, the procedures for the correction of the effects of the main error sources and for the stress uncertainty estimation, are proposed. The practical application of such procedures allow the user to highlight the relative magnitude of the error and stress uncertainty associated with the main influence parameters

Multifunctional crop trait ontology for breeders' data: field book, annotation, data discovery and semantic enrichment of the literature

The ‘Crop Ontology’ database we describe provides a controlled vocabulary for several economically important crops. It facilitates data integration and discovery from global databases and digital literature. This allows researchers to exploit comparative phenotypic and genotypic information of crops to elucidate functional aspects of traits

TILLING - a shortcut in functional genomics

Recent advances in large-scale genome sequencing projects have opened up new possibilities for the application of conventional mutation techniques in not only forward but also reverse genetics strategies. TILLING (Targeting Induced Local Lesions IN Genomes) was developed a decade ago as an alternative to insertional mutagenesis. It takes advantage of classical mutagenesis, sequence availability and high-throughput screening for nucleotide polymorphisms in a targeted sequence. The main advantage of TILLING as a reverse genetics strategy is that it can be applied to any species, regardless of its genome size and ploidy level. The TILLING protocol provides a high frequency of point mutations distributed randomly in the genome. The great mutagenic potential of chemical agents to generate a high rate of nucleotide substitutions has been proven by the high density of mutations reported for TILLING populations in various plant species. For most of them, the analysis of several genes revealed 1 mutation/200–500 kb screened and much higher densities were observed for polyploid species, such as wheat. High-throughput TILLING permits the rapid and low-cost discovery of new alleles that are induced in plants. Several research centres have established a TILLING public service for various plant species. The recent trends in TILLING procedures rely on the diversification of bioinformatic tools, new methods of mutation detection, including mismatch-specific and sensitive endonucleases, but also various alternatives for LI-COR screening and single nucleotide polymorphism (SNP) discovery using next-generation sequencing technologies. The TILLING strategy has found numerous applications in functional genomics. Additionally, wide applications of this throughput method in basic and applied research have already been implemented through modifications of the original TILLING strategy, such as Ecotilling or Deletion TILLING

An Induced Mutation in Tomato eIF4E Leads to Immunity to Two Potyviruses

BACKGROUND: The characterization of natural recessive resistance genes and Arabidopsis virus-resistant mutants have implicated translation initiation factors of the eIF4E and eIF4G families as susceptibility factors required for virus infection and resistance function. METHODOLOGY/PRINCIPAL FINDINGS: To investigate further the role of translation initiation factors in virus resistance we set up a TILLING platform in tomato, cloned genes encoding for translation initiation factors eIF4E and eIF4G and screened for induced mutations that lead to virus resistance. A splicing mutant of the eukaryotic translation initiation factor, S.l_eIF4E1 G1485A, was identified and characterized with respect to cap binding activity and resistance spectrum. Molecular analysis of the transcript of the mutant form showed that both the second and the third exons were miss-spliced, leading to a truncated mRNA. The resulting truncated eIF4E1 protein is also impaired in cap-binding activity. The mutant line had no growth defect, likely because of functional redundancy with others eIF4E isoforms. When infected with different potyviruses, the mutant line was immune to two strains of Potato virus Y and Pepper mottle virus and susceptible to Tobacco each virus. CONCLUSIONS/SIGNIFICANCE: Mutation analysis of translation initiation factors shows that translation initiation factors of the eIF4E family are determinants of plant susceptibility to RNA viruses and viruses have adopted strategies to use different isoforms. This work also demonstrates the effectiveness of TILLING as a reverse genetics tool to improve crop species. We have also developed a complete tool that can be used for both forward and reverse genetics in tomato, for both basic science and crop improvement. By opening it to the community, we hope to fulfill the expectations of both crop breeders and scientists who are using tomato as their model of study

Assessing the genetic variation of Ty-1 and Ty-3 alleles conferring resistance to Tomato Yellow Leaf Curl Virus in a broad tomato germplasm

The online version of this article (doi:10.1007/s11032-015-0329-y) contains supplementary material, which is available to authorized users.[EN] Tomato yellow leaf curl virus (TYLCV) hampers tomato production worldwide. Our previous studies have focussed on mapping and ultimately cloning of the TYLCV resistance genes Ty-1 and Ty-3. Both genes are derived from Solanum chilense and were shown to be allelic. They code for an RNA-dependent RNA polymerase (RDR) belonging to the RDR gamma type defined by a DFDGD catalytic domain. In this study, we first fine-mapped the TYLCV resistance in S. chilense LA1932, LA1960 and LA1971. Results showed that chromosomal intervals of the causal genes in these TYLCV-resistant accessions overlap and cover the region where Ty-1/Ty-3 is located. Further, virus-induced gene silencing was used to silence Ty-1/Ty-3 in tomato lines carrying TYLCV resistance introgressed from S. chilense LA1932, LA1938 and LA1971. Results showed that silencing Ty-1/Ty-3 compromised the resistance in lines derived from S. chilense LA1932 and LA1938. The LA1971-derived material remained resistant upon silencing Ty-1/Ty-3. Further, we studied the allelic variation of the Ty-1/Ty-3 gene by examining cDNA sequences from nine S. chilense-derived lines/accessions and more than 80 tomato cultivars, landraces and accessions of related wild species. The DFDGD catalytic domain of the Ty-1/Ty-3 gene is conserved among all tomato lines and species analysed. In addition, the 12 base pair insertion at the 5-prime part of the Ty-1/Ty-3 gene was found not to be specific for the TYLCV resistance allele. However, compared with the susceptible ty-1 allele, the Ty-1/Ty-3 allele is characterized by three specific amino acids shared by seven TYLCV-resistant S. chilense accessions or derived lines. Thus, Ty-1/Ty-3-specific markers can be developed based on these polymorphisms. Elevated transcript levels were observed for all tested S. chilense RDR alleles (both Ty-1 and ty-1 alleles), demonstrating that elevated expression level is not a good selection criterion for a functional Ty-1/Ty-3 allele.The infectious TYLCV clone was kindly provided by Professor Eduardo Rodriguez Bejarano (Universidad de Malaga). We thank Dick Lohuis for his help with agro-infiltrations, Marc Hendriks and Marjon Arens for RNA isolation and sequencing. This project was financed by the Centre for BioSystems Genomics (CBSG), which is part of the Netherlands Genomics Initiative/Netherlands Organization for Scientific Research (http://www.cbsg.nl). Olga Julian was granted a scholarship by Generalitat Valenciana. 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AgBioData consortium recommendations for sustainable genomics and genetics databases for agriculture

The future of agricultural research depends on data. The sheer volume of agricultural biological data being produced today makes excellent data management essential. Governmental agencies, publishers and science funders require data management plans for publicly funded research. Furthermore, the value of data increases exponentially when they are properly stored, described, integrated and shared, so that they can be easily utilized in future analyses. AgBioData (https://www.agbiodata.org) is a consortium of people working at agricultural biological databases, data archives and knowledgbases who strive to identify common issues in database development, curation and management, with the goal of creating database products that are more Findable, Accessible, Interoperable and Reusable. We strive to promote authentic, detailed, accurate and explicit communication between all parties involved in scientific data. As a step toward this goal, we present the current state of biocuration, ontologies, metadata and persistence, database platforms, programmatic (machine) access to data, communication and sustainability with regard to data curation. Each section describes challenges and opportunities for these topics, along with recommendations and best practices

Rapid identification of causal mutations in tomato EMS populations via mapping-by-sequencing

The tomato is the model species of choice for fleshy fruit development and for the Solanaceae family. Ethyl methanesulfonate (EMS) mutants of tomato have already proven their utility for analysis of gene function in plants, leading to improved breeding stocks and superior tomato varieties. However, until recently, the identification of causal mutations that underlie particular phenotypes has been a very lengthy task that many laboratories could not afford because of spatial and technical limitations. Here, we describe a simple protocol for identifying causal mutations in tomato using a mapping-by-sequencing strategy. Plants displaying phenotypes of interest are first isolated by screening an EMS mutant collection generated in the miniature cultivar Micro-Tom. A recombinant F2 population is then produced by crossing the mutant with a wild-type (WT; non-mutagenized) genotype, and F2 segregants displaying the same phenotype are subsequently pooled. Finally, whole-genome sequencing and analysis of allele distributions in the pools allow for the identification of the causal mutation. The whole process, from the isolation of the tomato mutant to the identification of the causal mutation, takes 6-12 months. This strategy overcomes many previous limitations, is simple to use and can be applied in most laboratories with limited facilities for plant culture and genotyping