Project description:The aim of the present work is to apply microarrays to identify candidate genes associated with the strategies adopted by different Solanum wild species to maintain their metabolism and limit the harm under water stress conditions. Wild genotypes are a rich source of novel genes for introduction into cultivated species to enhance stress tolerance. The mechanisms in which genes act can involve different pathways inside the cell, resulting in the expression of stress responsive genes. We will identify genes whose expression change in the leaves of potato plants subjected to drought stress. We will analyze different Solanum genotypes in six moments of water stress (0,1,3,5,7 & 10 days after initiation of stress). We will selected 4 time-points (days 0,5,7 & 10) and 2 genotypes (with divergence behavior) for microarrays analysis. The candidate genes obtained with this approach will be validated by semi-quantitative northern blot analysis (Typhoon scanner) and/or RT-PCR in each time-point of treatment and with all wild genotypes in study. This will help us to get a better understanding respect the dynamics of expression of a great number of genes in response to stress, as well as to establish functional relations between the involved genes. Research Plan: 5-weeks-old Solanum wild species plants were randomly sorted in 2 groups (25 plants per group). One group was submitted to non-irrigated conditions, withholding water and the other was normally watered. For microarray analysis, leaflets tissue from 4 plants per group of 4 time-points (days 0,5,7, & 10) were pooled, frozen in liquid nitrogen and stored at –80°C.RNA of the pool was extracted following TRIZOL method (Invitrogen). RNA integrity was verified on agarose gel. Hybridizations were performed with n genotypes,t time-points and r repeats. The hybridizations proposed in this experiment to enhance statistical robustness of data will be: 32 slides = n*t*r ; (2 genotypes; 4 time-points; 4 repeats (dye-swap). Keywords: Direct comparison
Project description:Transcriptomics study which main goal is to elucidate the programme of gene expression triggered by water stress in leaflets of the drought-tolerant wild-related tomato Solanum pennellii (acc. PE47) compared with domesticated tomato (S. lycopersicum, cv. P73). In this study we used S. lycopersicum (Sl) (cv. P73) and S. pennellii (Sp) (acc. PE47) species displaying remarkable divergences regarding drought tolerance, to investigate the physiological and molecular responses in leaves of plants grown without stress (control) and after four days of water withholding (water stress, WS), when plant water loss was significant but leaves did not show visual dehydration symptoms yet. Significant physiological differences between species were found, showing Sp leaves higher ability to avoid water loss. Leaf transcriptomic analysis showed important constitutive expression differences between Sp and Sl, including genes with unknown function. In relation to the genes specifically induced by drought in Sp, those linked to stomatal closure, cell wall and primary carbohydrate metabolism and, specially, nitrogen metabolism were identified. Thus, genes linked to NH4+ assimilation, GOGAT/GS cycle and the GDH- and GABA-shunt were specifically induced by water stress in leaves of Sp. Our results showed also the up-regulation in Sp of genes involved in JA biosynthesis pathway, which were induced in both conditions, whereas genes involved in ET biosynthesis were specifically induced under WS. Regarding ET signaling, ERF genes were up-regulated by WS in Sp, hinting at the importance of these transcriptional regulators in the drought response of Sp.
Project description:The potato is susceptible to water stress at all stages of development. We examined four clones of tetraploid potato, Cardinal, Desirée, Clone 37 FB and Mije, from the germplasm bank of the National Institute of Agricultural Research (INIA) in Chile. Water stress was applied by suspending irrigation at the beginning of tuberization. Stomatal conductance, tuber and plant fresh and dry weight was used to categorize water stress tolerance. Cardinal had high susceptibility to water stress. Desirée was less suscepetible than Cardinal and had some characteristics of tolerance. Mije had moderate and Clon 37 FB high tolerance. Differential gene expression in leaves from plants with and without water stress were examined using transcriptome sequencing. Water stress susceptible Cardinal had the fewest differentially expressed genes at 101, compared to Desirée at 1867, Clon 37 FB at 1179 and Mije at 1010. Water stress tolerance was associated with up-regulation of expression of transcription factor genes and genes involved in osmolyte and polyamine biosynthesis. Increased expression of genes encoding late embryogenesis abundant (LEA) and dehydrin proteins along with decreased expression of genes involved in nitrate assimilation and amino acid metabolism were found for clones showing water stress tolerance. The results also show that water deficit was associated with reduced biotic stress responses. Additionally, heat shock protein genes were differentially expressed in all clones except for highly susceptible Cardinal. Together the gene expression study demonstrates variation in the molecular pathways and biological processes in response to water stress contributing to tolerance and susceptibility.
Project description:Understanding the genetic basis of plants’ response to environmental stresses such as drought and salinity is vital for improving the future crop productivity and for deciphering the evolutionary mechanisms of adaptation and speciation. Here, we screened for genes and functional groups that are potentially involved in drought tolerance in tomato by comparing genome-wide transcriptome profiles of drought-sensitive S. lycopersicum and drought-tolerant S. pimpinellifolium populations under control and water deficit conditions. We also compared the transcriptome profiles from this study and a previous salt treatment study to investigate expression similarities and differences in gene expression patterns between water and salt stress responses, which are physiologically and biochemically similar. Stress-induced genes such as dehydration responsive element binding (DREB) protein, ABA-response element binding factor (AREB)-like protein, heat shock proteins, and chaperones were commonly up-regulated in S. lycopersicum and S. pimpinellifolium. Genes such as WRKY transcription factors and 1-aminocyclopropane-1-carboxylate (ACC) synthase exhibited striking differences in both the baseline expression under the control condition as well as expression changes in response to water deficit, suggesting that the two species have accumulated heritable differences in gene expression patterns. At the genome scale, there was a tendency that down-regulated genes in S. lycopersicum are more neutral or even up-regulated in S. pimpinellifolium, suggesting that S. pimpinellifolium may be able to maintain cellular activities during prolonged droughts. In comparison of water and salt stress responses, known stress-induced genes such as DREB protein, AREB-like protein, and nine-cis-epoxycarotenoid dioxygenase (NCED) were commonly up-regulated in response to these stresses. However, we also found fundamental differences between these stress responses in terms of genome-wide expression patterns, partly attributable to the difference in how these stresses were applied during the experiments.
Project description:Understanding the genetic basis of plants’ response to environmental stresses such as drought and salinity is vital for improving the future crop productivity and for deciphering the evolutionary mechanisms of adaptation and speciation. Here, we screened for genes and functional groups that are potentially involved in drought tolerance in tomato by comparing genome-wide transcriptome profiles of drought-sensitive S. lycopersicum and drought-tolerant S. pimpinellifolium populations under control and water deficit conditions. We also compared the transcriptome profiles from this study and a previous salt treatment study to investigate expression similarities and differences in gene expression patterns between water and salt stress responses, which are physiologically and biochemically similar. Stress-induced genes such as dehydration responsive element binding (DREB) protein, ABA-response element binding factor (AREB)-like protein, heat shock proteins, and chaperones were commonly up-regulated in S. lycopersicum and S. pimpinellifolium. Genes such as WRKY transcription factors and 1-aminocyclopropane-1-carboxylate (ACC) synthase exhibited striking differences in both the baseline expression under the control condition as well as expression changes in response to water deficit, suggesting that the two species have accumulated heritable differences in gene expression patterns. At the genome scale, there was a tendency that down-regulated genes in S. lycopersicum are more neutral or even up-regulated in S. pimpinellifolium, suggesting that S. pimpinellifolium may be able to maintain cellular activities during prolonged droughts. In comparison of water and salt stress responses, known stress-induced genes such as DREB protein, AREB-like protein, and nine-cis-epoxycarotenoid dioxygenase (NCED) were commonly up-regulated in response to these stresses. However, we also found fundamental differences between these stress responses in terms of genome-wide expression patterns, partly attributable to the difference in how these stresses were applied during the experiments. We compared Affymetrix microarray transcriptome profiles of root tissues from three natural populations each of S. lycopersicum and S. pimpinellifolium (3-4 individuals in each population as biological replicates under control (well-watered) and water deficit treatments.