Root transcriptional responses of two melon (Cucumis melo) genotypes with contrasting tolerance to Monosporascus cannonballus infection
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ABSTRACT: We used a melon oligo-based microarray to investigate the gene expression responses of two melon genotypes with contrasting resistance to Monosporascus cannonballus at 1 and 3 days after infection
Project description:Melon (Cucumis melo L.) is a commercially important fruit crop that is cultivated worldwide. The melon research community has recently benefited from the determination of a complete draft genome sequence and the development of associated genomic tools, which have allowed us to focus on small RNAs (sRNAs). These are short, non-coding RNAs 21â24 nucleotides in length with diverse physiological roles. In plants, they regulate gene expression and heterochromatin assembly, and control protection against virus infection. Much remains to be learned about the role of sRNAs in melon. We constructed 10 sRNA libraries from two stages of developing ovaries, fruits and photosynthetic cotyledons infected with viruses, and carried out high-throughput pyrosequencing. We catalogued and analyzed the melon sRNAs, resulting in the identification of 26 known miRNA families (many conserved with other species), the prediction of 84 melon-specific miRNA candidates, the identification of trans-acting siRNAs, and the identification of chloroplast, mitochondrion and transposon-derived sRNAs. In silico analysis revealed more than 400 potential targets for the conserved and novel miRNAs. This analysis provides insight into the composition and function of the melon small RNAome, and paves the way towards an understanding of sRNA-mediated processes that regulate melon fruit development and melonâvirus interactions. 11 small RNA libraries from several tissues of melon are included en the raw data. 2 samples from ovary, 2 samples from fruit, 1 sample from healthy cotyledons (Cultivar Tendral), 1 samples from healthy cotyledons (genotype TGR-1551), 1 sample from cotyledons (cultivar Tendral) infected with Watermelon mosaic virus (WMV), 1 sample from cotyledons (cultivar TGR-1551) infected with WMV, 1 sample from cotyledons (cultivar Tendral) infected with Melon necrotic spot virus (MNSV, Malfa5 isolate), 1 sample from cotyledons (cultivar Tendral) infected with MNSV (chimeric virus with Malfa5-264 isolates), 1 library from synthetic RNA oligos. Raw reads were obtained from two independent 454 runs, ~22,000 reads each one, to a total of 447,180 reads
Project description:Melon (Cucumis melo L.) is a commercially important fruit crop that is cultivated worldwide. The melon research community has recently benefited from the determination of a complete draft genome sequence and the development of associated genomic tools, which have allowed us to focus on small RNAs (sRNAs). These are short, non-coding RNAs 21â24 nucleotides in length with diverse physiological roles. In plants, they regulate gene expression and heterochromatin assembly, and control protection against virus infection. Much remains to be learned about the role of sRNAs in melon. We constructed 10 sRNA libraries from two stages of developing ovaries, fruits and photosynthetic cotyledons infected with viruses, and carried out high-throughput pyrosequencing. We catalogued and analyzed the melon sRNAs, resulting in the identification of 26 known miRNA families (many conserved with other species), the prediction of 84 melon-specific miRNA candidates, the identification of trans-acting siRNAs, and the identification of chloroplast, mitochondrion and transposon-derived sRNAs. In silico analysis revealed more than 400 potential targets for the conserved and novel miRNAs. This analysis provides insight into the composition and function of the melon small RNAome, and paves the way towards an understanding of sRNA-mediated processes that regulate melon fruit development and melonâvirus interactions.
Project description:RNA-Seq was conducted among sergeant bulks of four sex types of melon flowers, namely monoecious (AAGG), gynoecious (AAgg), hermaphrodite (aaGG), and andromonoecious (aagg), a total of about 105 million reads were generated from the melon transcriptome using Solexa sequencing.Totally 79,698 unigenes were generated and 75,537 unigenes were mapped to 11,805 annotated proteins in assembled melon genome (Garcia-Mas et al., 2012). Transcripts related to photomorphogenesis and flower development in plants were found, Most of the genes encoding plant hormone metabolism related protein, others related to flora development including Tasselseeds and male sterility genes which in phytohormones pathway were also detected. Comparison each two bulks (AAGG:AAgg, AAGG:aaGG, aagg:AAgg and aaGG:aagg ) exhibited different profiles of putative genes (include 745, 1342, 858 and 571 different expression genes, respectively). mRNA profiles of four sex types of melon flowers, namely monoecious (AAGG), gynoecious (AAgg), hermaphrodite (aaGG), and andromonoecious (aagg) were were generated by deep sequencing using Illumina Hiseq 2000.
Project description:Investigation gene expression level changes in four different melon fruits at four different developmental satges Transcriptomic analysis of developing melon fruits from two climacteric (cvs. Védrantais and Dulce) and two non-climacteric (cv. Piel de sapo and accession PI 161375) at 15, 25, 35 Days After Pollination (DAP) and Harvest stage
Project description:Filamentous fungi including mushrooms frequently and spontaneously degenerate during subsequent culture maintenance on artificial media, which shows the loss or reduction abilities of asexual sporulation, sexuality, fruiting and production of secondary metabolites, thus leading to economic losses during mass production. To better understand the underlying mechanisms of fungal degeneration, the model fungus Aspergillus nidulans was employed in this study for comprehensive analyses. First, linkage of oxidative stress to culture degeneration was evident in A. nidulans. Taken together with the verifications of cell biology and biochemical data, a comparative mitochondrial proteome analysis revealed that, unlike the healthy wild type, a spontaneous fluffy sector culture of A. nidulans demonstrated the characteristics of mitochondrial dysfunctions. Relative to the wild type, the features of cytochrome c release, calcium overload and up-regulation of apoptosis inducing factors evident in sector mitochondria suggested a linkage of fungal degeneration to cell apoptosis. However, the sector culture could still be maintained for generations without the signs of growth arrest. Up-regulation of the heat shock protein chaperones, anti-apoptotic factors and DNA repair proteins in the sector could account for the compromise in cell death. The results of this study not only shed new lights on the mechanisms of spontaneous degeneration of fungal cultures but will also provide alternative biomarkers to monitor fungal culture degeneration. Label-free quantitative proteomic analysis of the WT and Sec mt proteins was performed as described previously. Briefly, purified mitochondrial proteins (100 ug) were diluted in the lysis buffer to a concentration of 5 g/ul and carboxyamidomethylated in 50 mM iodoacetamide for 40 min at room temperature in the dark. The proteins were digested with an endoprotease Lys-C (Roche Applied Science, Indianapolis, IN) at a final substrate/enzyme ratio of 100:1 (w/w) at 37 C for 3 h. The Lys-C digests were further treated with modified sequencing grade trypsin (Roche Applied Science, Indianapolis, IN) at a final substrate/enzyme ratio of 50:1 (w/w) at 37 C for 20 h. After digestion, the peptide mixture was passed through an ultra-filter unit (Millipore, Billerica, MA) with a molecular weight cut-off of 10 kDa and acidified by formic acid (0.1% final concentration) for mass spectrum analysis. A linear ion trap-orbitrap liquid chromatography-tandem mass spectrometry system (LTQ-Orbitrap, Thermo Fisher Scientific, San Josse, CA) equipped with a nanospray ion source was used for full MS scan analysis followed by five MS/MS scans in the LTQ on the five most intense ions from the MS spectrum. Three parallel runs were performed consecutively for each sample. The software DeCyder MS (ver 2.0) (GE Healthcare, Pittsburgh, PA) was used to generate the peak lists from all the runs and the data were then automatically searched using the SEQUEST (ver. 2.7) (Thermo Fisher Scientific, San Josse, CA) against the A. nidulans genome archive (ver. S03-M05-R01, containing 10,644 protein entries) at AspGD database (www.aspgd.org) with the following parameters: peptide mass tolerance set to 10 ppm; fragment tolerance set to 0.8 Da, and two missed trypsin cleavages. Carbamidomethylation of cysteine was searched as a fixed modification, whereas N-acetyl protein and oxidation of methionine were searched as variable modifications. For protein identification, all peptide matches were filtered by a maximum false discovery rate (FDR) index less than 0.01, delta Cn larger than 0.1 and Xcorr scores of greater than 1.7, 2.0 and 3.0 for +1, +2, and +3 charged ions, respectively. The peptides were discarded for further analysis if mapped to more than two different proteins. A protein was considered identifiable with at least one unique peptide detected for more than twice in the three replicates of each sample. The MS data have been deposited in the PRIDE proteomics identification database under accession numbers ? List of the identified peptides and matched proteins is provided in Supplemental Table 1. Quantification of proteins was performed by estimating the normalized spectral index (SIN) for each protein, which combines the features of peptide count, spectral count, fragment-ion intensity and protein length as described by Griffin et al.. Reliability of technical repeats was analyzed by calculating the multivariate Pearson correlation coefficients using the software SigmaPlot (ver. 8.0). FDR estimation of differentially expressed proteins was conducted using a mixture model-based method. Heat mapping analysis was conducted using the program Matlab (R2009a). The significance of differentially expressed proteins between the samples was tested with the cutoff values of P less than or equal to 0.05 and FDR less than or equal to 0.05.
Project description:RNA-Seq was conducted among sergeant bulks of four sex types of melon flowers, namely monoecious (AAGG), gynoecious (AAgg), hermaphrodite (aaGG), and andromonoecious (aagg), a total of about 105 million reads were generated from the melon transcriptome using Solexa sequencing.Totally 79,698 unigenes were generated and 75,537 unigenes were mapped to 11,805 annotated proteins in assembled melon genome (Garcia-Mas et al., 2012). Transcripts related to photomorphogenesis and flower development in plants were found, Most of the genes encoding plant hormone metabolism related protein, others related to flora development including Tasselseeds and male sterility genes which in phytohormones pathway were also detected. Comparison each two bulks (AAGG:AAgg, AAGG:aaGG, aagg:AAgg and aaGG:aagg ) exhibited different profiles of putative genes (include 745, 1342, 858 and 571 different expression genes, respectively).