Transcriptome profiling and expression analyses of genes critical to wheat adaptation to low temperature
ABSTRACT: Understanding the mechanism of low temperature (LT) adaptation is crucial to the development of cold-tolerant crops. To identify the genes involved in the development of LT tolerance in the crown of hexaploid wheat we examined the global changes in genes expression during cold-treatment using the Affymetrix Wheat Genome Chip. Overall design: Time-series experiment with 4 genotypes x 8 time-points X 3 biological replicates in random block design; 96 hybridizations
Project description:Understanding the mechanism of low temperature (LT) adaptation is crucial to the development of cold-tolerant crops. To identify the genes involved in the development of LT tolerance in the crown of hexaploid wheat we examined the global changes in genes expression during cold-treatment using the Affymetrix Wheat Genome Chip. Time-series experiment with 4 genotypes x 8 time-points X 3 biological replicates in random block design; 96 hybridizations
Project description:We conducted microarray analysis to study comprehensive changes of gene expression profile under long-term low-temperature (LT) treatment and to identify other LT-responsive genes related with cold acclimation in seedling leaves and crown tissues (shoots containing apical meristems) of a synthetic hexaploid wheat line. The microarray analysis revealed marked up-regulation of a number of Cor/Lea genes and fructan biosynthesis-related genes under the long-term LT treatment. For validation of the microarray data, we selected four synthetic wheat lines, which contained the A and B genomes from a tetraploid wheat cultivar Langdon and the diverse D genomes originating from the different Ae. tauschii accessions, with distinct levels of freezing tolerance after cold acclimation. Quantitative RT-PCR analyses showed that the transcription accumulated levels of the Cor/Lea, CBF, and fructan biosynthesis-related genes were higher in more freezing-tolerant lines than those in the sensitive lines. The fructan biosynthesis pathway would be associated with cold acclimation to develop wheat freezing tolerance and related with diversity of the freezing tolerance level in addition to the CBF-mediated Cor/Lea expression pathway. Expression patterns were compared between a synthetic wheat line which treated 24℃ and 4℃. Total RNA samples were respectively isolated from leaves and crown tissues of the synthetic line grown at normal temperature for 3 weeks and then at 4°C for 12 and 6 weeks. Two independent experiments were conducted in each exprement.
Project description:We have employed whole genome microarray expression profiling as a discovery platform to identify genes to alter the transcript accumulation levels in a grass-clump dwarf line, which is a synthetic hexaploid line from triploid hybrids crossed between tetraploid wheat (Triticum turgidum ssp. durum cv. Langdon) and a diploid wheat relative Aegilops umbellulata (KU-4052). Up-regulation of metabolic and catabolic processes-related genes for cell wall-associated molecules was observed, and down-regulation of wheat APETALA1-like MADS-box genes, considered to act as flowering promoters, was found in the grass-clump dwarf line. Unusual expression of the branching-related SPLs and flowering time regulation-related MADS-box genes could explain the grass-clump dwarf phenotype. Overall design: Expression patterns were compared between the two synthetic hexaploid lines showing the wild-type phenotype (as a reference) and grass-clump dwarf. Total RNA samples were isolated from crown tissues of the plants grown at 24°C under long day (18-h light and 6-h dark) condition for 50 days. Two independent experiments were conducted in each exprement.
Project description:Two azide mutagenized lines Freeze Resistance (FR, 75% survival) and Freeze Susceptible (FS, 30% survival) were compared with and without 4°C ± 1.5 cold acclimation of crown tissue to identify genes responsible for the difference in freeze resistance. Keywords: Wheat cold acclimation, stress response, cold, low temperature Experiment design (8 hybridizations): Genotype: SD16029 (FR) or SD16169 (FS) Temperature: 25°C or 4°C
Project description:Gene expression levels of newly synthetic triploid wheat (ABD), its chromosome-doubled hexaploid (AABBDD), stable synthetic hexaploid (AABBDD), and their parents, Triticum turgidum (accession KU124, AABB) and Aegilops tauschii (accession KU2074, DD) were compared to understand genome-wide change of gene expressions during the course of amphidiploidization and genome stabilization. Stable synthetic hexaploid which were maintained through self-pollinations for 13 generations using the same combinations of the parents for production of synthetic common wheat. Overall design: Amphidiploidization event between T. turgidum ssp. dicoccum (AABB) and Ae. tauschii ssp. strangulata (DD) was recreated. Gene expression levels of newly synthetic triploid wheat (ABD), its chromosome-doubled hexaploid (AABBDD), stable synthetic hexaploid (AABBDD), and their parents, Triticum turgidum (AABB) and Aegilops tauschii (DD) were compared. Total RNA of each line was extracted from three biological replicates of two leaves seedlings.
Project description:We have employed whole genome microarray expression profiling as a discovery platform to identify genes to alter the transcript accumulation levels in grass-clump dwarf lines, which are synthetic hexaploid lines from triploid hybrids crossed between tetraploid wheat (Triticum turgidum ssp. durum cv. Langdon or T. turgidum ssp. carthlicum) and diploid wheat progenitor Aegilops tauschii (KU2025). No up-regulation of defense-related genes was observed under the normal temperature, and down-regulation of wheat APETALA1-like MADS-box genes, considered to act as flowering promoters, was found in the grass-clump dwarf lines. Together with small RNA sequencing analysis of the grass-clump dwarf line, unusual expression of the miR156/SPLs module could explain the grass-clump dwarf phenotype. Expression patterns were compared between the three synthetic hexaploid lines showing the wild-type phenotype (as a reference) and grass-clump dwarf. Total RNA samples were isolated from crown tissues of the plants grown at 24°C under long day (18-h light and 6-h dark) condition for 8 weeks. Two independent experiments were conducted in each exprement.
Project description:Wheat is a cereal grain and one of the world’s major food crops. Recent advances in wheat genome sequencing are by now facilitating genomic and proteomic analyses of this crop. However, little is known about the protein levels of hexaploid versus tetraploid wheat cultivars, and knowledge on phosphorylated proteins still limited. Using our recently established (phospho)proteomic workflow, we performed a parallel analysis of the proteome and phosphoproteome on seedling leaves from two hexaploid wheat cultivars (Pavon 76 and USU-Apogee) and a tetraploid wheat (Senatore Cappelli). This revealed that a large portion of proteins and phosphosites can be quantified in all cultivars. Our shotgun proteomics data revealed a high similarity between hexaploid and tetraploid varieties with respect to protein abundance. However, we could identify a set of proteins that were differentially abundant between hexaploid and tetraploid cultivars. In addition, already at seedling stage, a small set of proteins were differential between the small (USU-Apogee) and larger hexaploid wheat cultivar (Pavon 76), which could potentially act as growth predictors. Finally, the phosphosites identified in this study can be retrieved from the in-house developed plant PTM-Viewer (bioinformatics.psb.ugent.be/webtools/ptm_viewer/), making this the first repository for phosphorylated wheat proteins. This paves the way for further in depth, quantitative (phospho)proteome-wide differential analyses upon a specific trigger or environmental change.
Project description:affy_hexaploid_wheat - hexa - Changes upon polyploidization in hexaploid wheat - transcriptomic changes in synthetic hexaploid derived from a cross between tetraploid and natural diploid. Study aims to understand regulation of gene expression in synthetic and natural wheat allohexaploids (Triticum aestivum), that combines the AB genome of T. turgidum and the D genome of Aegilops tauschii; and which we have recently characterized as genetically stable. We conducted a comprehensive genome-wide analysis of gene expression that allowed us to compare the effect of variability of the D genome progenitor, the trans-generation stability as well as comparison to natural wheat allohexaploid. We used the Affymetrix GeneChip® Wheat Genome Array, on which 55,049 transcripts are represented. Additive expression was shown to represent majority of expression regulation in the synthetic allohexaploids, where expression for more than 93% of transcripts was equal to the one evaluated from equal mixture of parental RNA. This leaves ~2000 (~7%) transcripts, which expression was non-additive. No global gene expression bias or dominance towards any of the progenitor genomes was observed whereas high trans-generational stability and low effect of the D genome progenitor variability were revealed. Our study suggests that gene expression regulation in wheat allohexaploids is early established upon allohexaploidization and highly conserved over generations, as demonstrated by the high similarity of expression with natural wheat allohexaploids. Keywords: genotype and ecotype comparison Overall design: 18 arrays - wheat
Project description:Different wheat cultivars may be classified as either winter or spring varieties depending on whether they require exposure to an extended period of cold in order to become competent to flower. Using a growth regime that mimics the conditions that occur during a typical winter in Britain, we wished to survey the genes that are involved in phase transition as well as those involved in cold-acclimation. Keywords: Time course Overall design: We wished to study the profiles of expression of genes involved in both phase transition (vegetative to reproductive growth transition) and cold-acclimation. To that end we we exposed plants to a gradual, stepped decline in both temperature and light. We sampled plants at three time points (3 weeks post-germination, 5 weeks post germination and 9 weeks post germination). We took samples from two separate tissues (crown and leaf) to se whether responses were different. We used two biological reps for each time point and tissue. Control plants were exposed to a delined in day-length and light intensity, but not in temperature.
Project description:The crown is the critical region for survival of winter wheat exposed to low temperature stresses. When wheat is exposed to non-freezing low temperatures, they can increase their freezing tolerance (cold acclimation, ACC). Changes within the apoplast are thought to be crucial for acquisition of freezing tolerance. However, how individual tissues within the ccrown, namely the shoot apical meristem (SAM, responsible for new shoot growth) and vascular transition zone (VTZ, located at the base of the crown)enhance tolerance to freezing has not yet been characterized. In the present study, we conducted shotgun proteomic analysis of the apoplast fluid to investigate ACC-induced proteins in the SAM and VTZ.