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Figueiredo A.,University of Lisbon | Loureiro A.,CIFC Biotrop IICT Institute Investigacao Cientifica Tropical | Batista D.,CIFC Biotrop IICT Institute Investigacao Cientifica Tropical | Monteiro F.,University of Lisbon | And 4 more authors.
BMC Research Notes | Year: 2013

Background: Coffee production in Africa represents a significant share of the total export revenues and influences the lives of millions of people, yet severe socio-economic repercussions are annually felt in result of the overall losses caused by the coffee berry disease (CBD). This quarantine disease is caused by the fungus Colletotrichum kahawae Waller and Bridge, which remains one of the most devastating threats to Coffea arabica production in Africa at high altitude, and its dispersal to Latin America and Asia represents a serious concern. Understanding the molecular genetic basis of coffee resistance to this disease is of high priority to support breeding strategies. Selection and validation of suitable reference genes presenting stable expression in the system studied is the first step to engage studies of gene expression profiling. Results: In this study, a set of ten genes (S24, 14-3-3, RPL7, GAPDH, UBQ9, VATP16, SAND, UQCC, IDE and β-Tub9) was evaluated to identify reference genes during the first hours of interaction (12, 48 and 72 hpi) between resistant and susceptible coffee genotypes and C. kahawae. Three analyses were done for the selection of these genes considering the entire dataset and the two genotypes (resistant and susceptible), separately. The three statistical methods applied GeNorm, NormFinder, and BestKeeper, allowed identifying IDE as one of the most stable genes for all datasets analysed, and in contrast GADPH and UBQ9 as the least stable ones. In addition, the expression of two defense-related transcripts, encoding for a receptor like kinase and a pathogenesis related protein 10, were used to validate the reference genes selected. Conclusion: Taken together, our results provide guidelines for reference gene(s) selection towards a more accurate and widespread use of qPCR to study the interaction between Coffea spp. and C. kahawae. © 2013 Figueiredo et al.; licensee BioMed Central Ltd.

Pinheiro C.,Institute Tecnologia Quimica e Biologica | Pinheiro C.,New University of Lisbon | Guerra-Guimaraes L.,CIFC Biotrop IICT Institute Investigacao Cientifica Tropical | David T.S.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Vieira A.,CIFC Biotrop IICT Institute Investigacao Cientifica Tropical
Environmental and Experimental Botany | Year: 2014

Mediterranean woody species are vulnerable to multiple stresses that negatively affect plant survival and productivity. Drought and heat are increasing threats to the agricultural and forestry systems, making it urgently necessary to determine the mechanisms of plant adaptation and survival. Proteomics allows for the characterisation of a large number of proteins in a given tissue/organ, providing an integrated picture of the molecular events involved in stress responses. In this paper, we have evaluated the contribution of proteomics for the identification of stress-responsive proteins and tolerance/adaptation mechanisms in woody plants of agronomic importance in the Mediterranean basin. A systematic review was performed (Web of Knowledge, 5th March 2013) on the relevant genera for this region: Quercus sp., Pinus sp., Eucalyptus sp., Vitis sp., Olive sp., and Citrus sp. The term Rosaceae was also included in the search due to the relevance of fruit tree crops of this family in the Mediterranean region. This systematic review highlighted the lack of extensive and comprehensive proteomic analyses for Mediterranean plants under stress. The approach retrieved 19 and 38 papers concerning the assessment of abiotic and biotic stresses, respectively, at the proteome level, and 20 and 46 papers, respectively, concerning analyses at the transcriptomics level. With regard to abiotic stress, gel-based proteomic methodologies (15 papers) enabled the identification and quantification of 395 stress-responsive proteins. These results revealed metabolic adjustments to stress, with major alterations in carbon, nitrogen, and amino acid metabolisms. The most consistently represented stress-responsive proteins were RuBisCO, RuBisCO activase, heat shock proteins, chlorophyll a/. b binding protein, and proteins from the oxygen-evolving complex. We concluded that gel-based proteomics revealed key proteins and metabolic pathways important for the ability of plants to adjustment to environmental fluctuations. The integration of this information with physiological, agronomic, and technological performance (e.g. survival, productivity, and food and technological quality) is essential for the sustainable development of the Mediterranean regions. © 2014 Elsevier B.V.

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