Project description:To improve our understanding of the organization and regulation of the wheat gene space, we established the first transcription map of a wheat chromosome (3B) by hybridizing the newly developed INRA GDEC Triticum aestivum NimbleGen 12x40k unigenes microarray with BAC pools from a new version of the 3B physical map as well as with cDNA probes from five tissues at three developmental stages each. By hybridizing the BAC pools with the wheat NimbleGen 40K unigenes chip we managed to map almost 3000 unigenes on the wheat chromosome 3B BACs and to study the organization of the wheat gene space along chromosome 3B. The sequences of the unigenes helped to perform functional and evolutionary analyses of these unigenes. By hybridizing the 15 cDNA samples from five organs at three developmental stages each we established the expression profiles of more than 32000 wheat unigenes. Particularly we focused on the expression of the unigenes mapped on wheat chromosome 3B to perform coexpression analyses.
Project description:The goal of this study was to calculate the degree of substantial equivalence between a GM potato line modified for late blight resistance and corresponding non-GM controls in the presence/absence of P. infestans inoculation. The GM phenotype was conferred through the introduction of the Rb gene which has been previously cloned from Solanum bulbocastanum (Song et al. 2003). The resulting GM phenotype was characterised by the expression of strong, partial resistance to P. infestans. By obtaining an expression profile from a GM and non-GM population before and after exposure to P. infestans, this effectively provides for the completion of two risk assessments: impact on the potato transcriptome of plus/minus genetic modification; before and after exposure to the pathogen. For each biological replicate, individuals from three separate GM lines (A, B, C) and three separate non-GM lines (D, E, F) were randomly sorted and cultivated for analysis. Each GM line contained an independent, single transgene insertion, which was verified through Southern Blot analysis. One day prior to pathogen exposure, the 4-week-old potato plants were transferred into a growth chamber with a 16-h light period, 90% relative humidity at a temperature of 15C. Plants were randomly selected and 14 individuals/line were inoculated with a field isolate of P. infestans. An additional 14 plants/line were sprayed with water. Inoculations were made with a sporangial suspension of 20,000/mL and plants were sprayed until run-off. Twenty-four hours post-inoculation, leaf tissue was collected from 9 plants per treatment, flash frozen in liquid nitrogen and pooled. Total leaf RNA was extracted using Trizol and the whole experiment was repeated three times (three biological replicates). Keywords: Direct comparison 19 hybs total
Project description:The transcriptome profile of arbuscular mycorrhiza established at 4 weeks post inoculation between Medicago truncatula and Glomus mosseae as well as between Medicago truncatula and Glomus intraradices is compared
Project description:Blackspot bruising is the tuber discoloration resulting from mechanical damage in susceptible crops and is a major quality and environmental problem for the potato industry. The present study is aimed at identifying differences in gene expression which are correlated with the development of bruise susceptibility or resistance in tubers. This may produce markers or diagnostic tests for use in predicting enhanced bruise susceptibility in field grown potato crops. Different potato cultivars were grown in field plots at ADAS Gleadthorpe, Nottinghamshire, UK during three growing seasons – 2003, 2004 and 2005. Random samples of tubers (8) were harvested by hand and cores excised from the subdermal tissues of the stolon end of the tubers and immediately frozen and stored in liquid nitrogen for RNA isolations. The remainder of the crops were stored at 4oC for bruise susceptibility testing - samples of tubers (30) from each crop were impacted and incubated for 48h at 30oC. Bruise susceptibility was scored, according to the intensity of pigmentation and the volume of tuber tissue affected, on a scale of 0 (very resistant) – 10 (very susceptible). Crops representing 'very susceptible', 'susceptible', 'moderate' and 'resistant' to bruising, were selected for RNA isolation. Total RNA’s were isolated from frozen cores by the TIGR hot phenol extraction method, precipitated by LiCl and treated with DNase. RNA concentrations were adjusted to ~2ug/ul and analysed on 1% TBE gels after formamide treatment. Any RNA’s which were not pure enough or showed degradation were discarded. Total RNA’s were pooled and 25ul samples aliquotted according to the required hybridisations indicated in the Excel spreadsheet and the on-line information form. Keywords: Direct comparison 32 hybs total
Project description:Genes specifically or induced in seed coat of Medicago truncatula 10-36 days after pollination were identified by comparing with genes expressed in other organs.
Project description:In order to understand effects of defective circadian clock on the global barley transcriptome, we analyzed the diel and circadian leaf transcriptomes in the barley cultivar Bowman and derived introgression lines carrying mutations in EARLY FLOWERING 3 (ELF3), LUX1, and EARLY MATURITY 7 (EAM7).
Project description:Heat stress is a major cause for yield loss in many crops, including vegetable crops. Even short waves of high temperature, becoming more frequent during recent years, can be detrimental. Pollen development is most heat-sensitive, being the main cause for reduced productivity under heat-stress across a wide range of crops. The molecular mechanisms involved in pollen heat-stress response and thermotolerance are however not fully understood. Recently, we have demonstrated that ethylene, a gaseous plant hormone, plays a role in tomato (Solanum lycopersicum) pollen thermotolerance. These results were substantiated in the current work showing that increasing ethylene levels by using an ethylene-releasing substance, ethephon, prior to heat-stress exposure, increased pollen quality. A proteomic approach was undertaken, to unravel the mechanisms underlying pollen heat-stress response and ethylene-mediated pollen thermotolerance in developing pollen grains. Proteins were extracted and analyzed by means of a gel LC-MS fractionation protocol, and a total of 1355 proteins were identified. A dataset of 721 proteins, detected in three biological replicates of at least one of the applied treatments, was used for all analyses. Quantitative analysis was performed based on peptide count. The analysis revealed that heat-stress affected the developmental program of pollen, including protein homeostasis (components of the translational and degradation machinery), carbohydrate and energy metabolism. Ethephon-pretreatment shifted the heat-stressed pollen proteome closer to the proteome under non-stressful conditions, namely, by showing higher abundance of proteins involved in protein synthesis, degradation, tricarboxylic acid cycle and RNA regulation. Furthermore, up-regulation of protective mechanisms against oxidative stress was observed following ethephon-treatment (including higher abundance of glutathione-disulfide reductase, glutaredoxin and protein disulfide isomerase). Taken together, the findings identified systemic and fundamental components of pollen thermotolerance, and serve as a valuable quantitative protein database for further research.
Project description:In present study, digital gene expression (DGE) profiling was performed to obtain a general picture of the transcriptomic implicated in the early development of R. venosa. Eighteen DGE libraries at six developmental stages of R. venosa were constructed, sequenced by IIIumina HiSeq 2500 platform. RNA from six developmental stages of R. venosa was sequenced using Illumina Hi-seq 2500. Each stage have three replicates