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      Comparative transcript profiling by SuperSAGE identifies novel candidate genes for controlling potato quantitative resistance to late blight not compromised by late maturity

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          Abstract

          Resistance to pathogens is essential for survival of wild and cultivated plants. Pathogen susceptibility causes major losses of crop yield and quality. Durable field resistance combined with high yield and other superior agronomic characters are therefore, important objectives in every crop breeding program. Precision and efficacy of resistance breeding can be enhanced by molecular diagnostic tools, which result from knowledge of the molecular basis of resistance and susceptibility. Breeding uses resistance conferred by single R genes and polygenic quantitative resistance. The latter is partial but considered more durable. Molecular mechanisms of plant pathogen interactions are elucidated mainly in experimental systems involving single R genes, whereas most genes important for quantitative resistance in crops like potato are unknown. Quantitative resistance of potato to Phytophthora infestans causing late blight is often compromised by late plant maturity, a negative agronomic character. Our objective was to identify candidate genes for quantitative resistance to late blight not compromised by late plant maturity. We used diagnostic DNA-markers to select plants with different field levels of maturity corrected resistance (MCR) to late blight and compared their leaf transcriptomes before and after infection with P. infestans using SuperSAGE (serial analysis of gene expression) technology and next generation sequencing. We identified 2034 transcripts up or down regulated upon infection, including a homolog of the kiwi fruit allergen kiwellin. 806 transcripts showed differential expression between groups of genotypes with contrasting MCR levels. The observed expression patterns suggest that MCR is in part controlled by differential transcript levels in uninfected plants. Functional annotation suggests that, besides biotic and abiotic stress responses, general cellular processes such as photosynthesis, protein biosynthesis, and degradation play a role in MCR.

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          Cross talk in defense signaling.

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            Selection of internal control genes for quantitative real-time RT-PCR studies during tomato development process

            Background The elucidation of gene expression patterns leads to a better understanding of biological processes. Real-time quantitative RT-PCR has become the standard method for in-depth studies of gene expression. A biologically meaningful reporting of target mRNA quantities requires accurate and reliable normalization in order to identify real gene-specific variation. The purpose of normalization is to control several variables such as different amounts and quality of starting material, variable enzymatic efficiencies of retrotranscription from RNA to cDNA, or differences between tissues or cells in overall transcriptional activity. The validity of a housekeeping gene as endogenous control relies on the stability of its expression level across the sample panel being analysed. In the present report we describe the first systematic evaluation of potential internal controls during tomato development process to identify which are the most reliable for transcript quantification by real-time RT-PCR. Results In this study, we assess the expression stability of 7 traditional and 4 novel housekeeping genes in a set of 27 samples representing different tissues and organs of tomato plants at different developmental stages. First, we designed, tested and optimized amplification primers for real-time RT-PCR. Then, expression data from each candidate gene were evaluated with three complementary approaches based on different statistical procedures. Our analysis suggests that SGN-U314153 (CAC), SGN-U321250 (TIP41), SGN-U346908 ("Expressed") and SGN-U316474 (SAND) genes provide superior transcript normalization in tomato development studies. We recommend different combinations of these exceptionally stable housekeeping genes for suited normalization of different developmental series, including the complete tomato development process. Conclusion This work constitutes the first effort for the selection of optimal endogenous controls for quantitative real-time RT-PCR studies of gene expression during tomato development process. From our study a tool-kit of control genes emerges that outperform the traditional genes in terms of expression stability.
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              Resistance gene-dependent plant defense responses.

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                Author and article information

                Journal
                Front Plant Sci
                Front Plant Sci
                Front. Plant Sci.
                Frontiers in Plant Science
                Frontiers Media S.A.
                1664-462X
                14 November 2013
                2013
                : 4
                : 423
                Affiliations
                [1] 1Department Plant Breeding and Genetics, Max Planck Institute for Plant Breeding Research Cologne, Germany
                [2] 2GenXPro GmbH Frankfurt, Germany
                [3] 3Saka-Pflanzenzucht GmbH & Co. KG Windeby, Germany
                Author notes

                Edited by: Ingo Schubert, Leibniz Institute of Plant Genetics and Crop Plant Research, Germany

                Reviewed by: Matthias Harbers, RIKEN Center for Life Science Technologies, Japan; Ryohei Terauchi, Iwate Biotechnology Research Center, Japan

                *Correspondence: Christiane Gebhardt, Department of Plant Breeding and Genetics, Max Planck Institute for Plant Breeding Research, Carl von Linné Weg 10, 50829 Cologne, Germany e-mail: gebhardt@ 123456mpipz.mpg.de

                This article was submitted to Plant Genetics and Genomics, a section of the journal Frontiers in Plant Science.

                †Present address: Astrid M. Draffehn, Life and Medical Sciences LIMES Institute Bonn, Genomics and Immunoregulation, Bonn, Germany;

                Li Li, State Key Laboratory of Tree Genetics and Breeding (Northeast Forestry University), Harbin, People's Republic of China

                Article
                10.3389/fpls.2013.00423
                3827546
                bab29687-336f-4dc2-8680-90ccb04adeb0
                Copyright © 2013 Draffehn, Li, Krezdorn, Ding, Lübeck, Strahwald, Muktar, Walkemeier, Rotter and Gebhardt.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 17 June 2013
                : 05 October 2013
                Page count
                Figures: 6, Tables: 4, Equations: 0, References: 90, Pages: 21, Words: 14498
                Categories
                Plant Science
                Original Research Article

                Plant science & Botany
                marker-assisted selection,late blight,transcript profiling,sage,potato
                Plant science & Botany
                marker-assisted selection, late blight, transcript profiling, sage, potato

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