Digital gene expression of WT and gi mutant under control and drought conditions in Arabidopsis
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ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE48810 | GEO | 2013/07/12
SECONDARY ACCESSION(S): PRJNA211822
REPOSITORIES: GEO
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Project description:We report the application of digital gene expression analysis for high-throughput profiling the different gene expression of WT and gi mutant under control and drought conditions. Examination gene expression of WT and gi under control and drought conditions
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Project description:Transcriptional reprogramming plays a key role in drought stress responses, preceding the onset of morphological and physiological adaptations. The best characterised signal regulating gene expression in response to drought is the phytohormone abscisic acid (ABA). ABA-regulated gene expression, biosynthesis and signalling are highly organised in a diurnal cycle, so that ABA-regulated physiological traits occur at the appropriate time of the day. The mechanisms that underpin such diel oscillations in ABA signals are poorly characterised. Here we uncover GIGANTEA (GI) as a key gate keeper of ABA-regulated transcriptional and physiological responses. Time-resolved RNAseq under different irrigation scenarios indicate that gi mutants produce an exaggerated ABA response at midday, despite accumulating wild-type levels ABA. The direct comparison with ABA-deficient mutants confirms the role of GI in controlling ABA-regulated genes and enabled us to distinguish a phase of high ABA sensitivity at dawn, characterized by low GI accumulation.
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Project description:Six months old seed grown under well-watered greenhouse conditions were used in the current study. The seedlings of the two wild emmer wheat genotypes were vernalized on a moist germination paper (Hofman Manufacturing, Inc, Jefferson, OR, USA) for three weeks in the dark at 4oC, followed by three days acclimation at 24oC. Seedlings were then transplanted into 5L pots containing a mixture of pure quartz sand and peat (4:1 v/v), supplemented with a slow release fertilizer (2 g/L, Osmocote® Standard 14-14-14, Scotts-Sierra Horticulture, Marysville, OH, USA) and by a weekly application of 100 ml/pot of 0.5X Murashigi and Skoog growth solution (Sigma Chemical Co., St Louis, MO, USA). Pots were placed in a screen-house under natural winter conditions (December to February; 5-18°C) in Haifa, Israel (Mt. Carmel; 35o01′ E, 32o45′ N; 480 m above sea level) for 10 weeks and irrigated daily by a drip irrigation system. Three weeks prior to application of terminal drought stress, the pots were transferred to a controlled environment greenhouse (22/18 oC; 12 h day/12 h night) in order to prevent rainfall during the drought experiment. Five pots per genotype served as biological replicates, plants of three pots of each genotype/treatment were used for transcriptome study whereas two additional pots were used for quantitative PCR analysis (altogether five biological replicated of were used for quantitative real time PCR). Three individual plants were grown in each pot, one plant was used for measurements of leaf relative water content (RWC) (Barrs and Weatherley 1968), whereas the other two plants were used for sampling of flag leaf tissue for RNA extraction. Terminal drought stress (D) was applied at inflorescence emergence stage (Zadoks 50-60), (Zadoks et al. 1974), after emergence of 1-2 spikes in all biological replicates of both genotypes. The time from transplanting to inflorescence emergence stage was not significantly different between genotypes (70 days in the S genotype and 72 days in the R genotype). Drought stress application initiated after irrigation with excess amount of water in order to assure that all pots start the experiment at the same soil water capacity. Drought stress was imposed by withholding water for eight days until stress symptoms (i.e. leaf rolling and wilting symptoms) were visible in plants of both genotypes. Stress symptoms were more visible in the S genotype, however, leaf relative water content (RWC), measured after eight days of drought stress was low but was not significantly different between the two genotypes (49.68%±1.48 in the R genotype and 53.34%±1.83 in the S genotype). The well-watered control (C) plants were irrigated daily by ample amount of water. Flag leaf tissues of drought-stressed plants and well-watered control plants were harvested, immediately frozen in liquid nitrogen, and stored at -80ºC for RNA extraction. ****[PLEXdb(http://www.plexdb.org) has submitted this series at GEO on behalf of the original contributor, Krugman Tamar. The equivalent experiment is TA33 at PLEXdb.] genotype: Drought resistant (R) Y12-3 - treated or untreated: Well-watered(3-replications); genotype: Drought resistant (R) Y12-3 - treated or untreated: terminal drought(3-replications); genotype: Drought susceptible (S) A24-39 - treated or untreated: Well-watered(3-replications); genotype: Drought susceptible (S) A24-39 - treated or untreated: terminal drought(3-replications)
2011-08-30 | E-GEOD-31762 | biostudies-arrayexpress
Project description:In rice, gene expression profiling under different stresses have been carried out earlier through microarrays and SAGE (Kawasaki et al., 2001; Rabbani et al., 2003,;. Yazaki et al., 2003, 2004; Walia et al., 2005; Zhou et al., 2005; Jiao et al., 2005; Zhou, 2007; Wang, et al., 2007)). The results reported so far were mostly derived from different stress treatments, array formats, species, tissue types, and time courses, which make it difficult for direct comparison among studies. Also, most of the stress experiments on expression profiles were conducted under laboratory, growth chamber or field grown plants subjected to stress in laboratory conditions, usually by inducing stress with exogenous compounds such as abscisic acid (ABA) or gibberellin (GA) genes (Yazaki et al., 2003, 2004; Rabbani et al., 2003), salt (Kawasaki et al., 2001; Rabbani et al., 2003; Walia et al., 2005), organ specific gene expression under high salinity (Jiao et al., 2005) and drought (Zhou et al., 2007), comparison with different cultivars (Wang et al., 2007) and other environmental stresses such as cold and drought (Rabbani et al., 2003; Zhou et al., 2005). However, such laboratory, growth chamber grown plants or field grown plants subjected to stress under laboratory conditions significantly differ from the field stress conditions and therefore the resultant plant response may not truly represent the actual stress conditions prevailing on field. This has been extensively reviewed by Mittler, 2006 and the need for utilizing data from multiple stress experiments for developing plants resistant to stress has been focused. The review also focuses on evaluating farmers experiences in stress management and kinds of stress that a plant is subjected prior to taking up research on stress resistances has been of significant importance. In the present study, to bridge the gap between the available data on drought stress and the gene regulation under field drought conditions has been paid attention. The outcome of this data will give new leads for breeders to design strategies to develop plants resistant to drought without much loss in yield. In the present study experiments were conducted under different field capacities using a rainout shelter to minimizing the effects of drought induction due to rains. The microarray analysis is thus expected to throw light on stress responsive gene expression in field drought conditions. Till now, to our knowledge there have been no reports on gene expression in rice under field drought conditions. Further, there is no data available on gene expression profiling during various stages of plant growth in leaf and panicle tissues under different levels of water stress. Here, we have focused on generating stress responsive gene (SRG) expression profiles in field drought conditions through the entire life cycle of rice plant. Optimal drought simulation was achieved by conducting a drought stress experiment and validated by a series of dry runs. The main reason for evaluating gene expression under drought in field condition is as drought itself is not limited to water stress but various other factors, such as heat, humidity, hot winds, and long day lengths effect the gene regulation. All these are likely to trigger common pathways that are also a part of drought stress response and therefore significantly affect the expression profiles. In the present study, field drought stress was initiated from 31 days after germination and transplantation to the field at 100%, 60%, 40% and 15% field capacities designated as A1, A2, A3, and A4. It took 7 days for the onset of the actual drought for the experimental plots to reach to their designated field capacities from 100% FC. Samples were collected from 7 days after drought stress to 33 days after drought stress to evaluate different drought stress regimes based on the developmental stages of the plant. At all the tested stress levels the data points were Hierarchically grouped and clustered sets of genes were examined and an approximate number of clusters to be formed were estimated. Finding a group of genes showing a similar pattern of gene expression by means of expression data from microarray analysis is the basis for identification of genes and pathways controlling a specific trait. We have followed unsupervised classification and clustered the genes hierarchically after normalizing expression data to obtain a set of co-regulated genes. Analysis of data revealed that there are at least 3 to 4 clusters which show differential regulation under each stress regime. The annotated genes falling in these clusters interestingly include a number of novel genes from N22 library along with some known candidate genes for drought tolerance which were earlier identified in our study (Markandeya et al., 2007). Expression pattern of these genes from 7 days after drought stress to 33 days after drought stress with their fold changes were evaluated in the present study. In particular, it is observed that many genes of unknown function are also co-regulated in an interesting pattern along with known genes.
2011-11-27 | GSE31974 | GEO