Project description:Our objective is to describe variations in expression profiles of transcriptional factors during endosperm development, especially during filling phase in which storage proteins are synthesized. Plants from cv. Recital were cultivated under controled conditions. Two biological repetitions were available for grain harvest. at 11 developmental stages (132, 172, 289, 323, 381, 437, 477, 527, 577, 635 and 686 degrees C day). RNA was extracted from all samples of seeds whose embryo was removed.cDNA synthesis, labelling, and hybridization were made according to manufacturer’s protocol (NimbleGen Arrays Users Guide). The SuperScript Double-Stranded cDNA synthesis kit (invitrogen) was used to transform RNA in double-stranded cDNA. The first strand was transcribed with an oligo dT primer and the SuperscriptTM II Reverse Transcriptase. The synthesis of second strand was performed according to the method of Gubler and Hoffman by using RNAse H, DNA ligase, DNA polymerase I. The cDNA was labeled with Cy3 by Random priming by using the NimbleGen One-Color DNA Labeling Kit. Each biological replicate including 11 samples was hybridized on independent microarray. A common sample was placed on both microarrays (stage 323 degrees C day, Biological Repetition 2). The Cy3-labeled cDNA samples were hybridized on microarray by using the NimbleGen Hybridization Kit at 42 degrees c for 18h. Microarrays were washed by using the components of the NimbleGen Wash Buffer Kit and NimbleGen Array Processing Accessories. Then, microarrays were scanned with an Innoscan 900 AL scanner (Innopsys) by using Mapix software. NimbleScan software was used to create signal intensities files from scanned images files. A grid was overlaid on the microarray image to identify the placement of the probes. The software extracts raw data and generates X,Y and signal reports providing the coordinates of each probe and its intensity
Project description:Investigation of genome wide expression level changes during 11 stages of wheat grain development in normal growth conditions (19°C).
Project description:For many potato cultivars, tuber yield is optimal at average day time temperatures in the range of 14-22 ⁰C. Further rises in ambient temperature can reduce or completely inhibit potato tuber production, with damaging consequences for both producer and consumer. In our previous work we observed that the steady-state expression level of the core circadian clock gene, TIMING OF CAB EXPRESSION 1 (TOC1), in potato tubers increased at moderately elevated temperature, whereas expression of the tuberisation signal gene StSP6A decreased along with tuber yield. In this study we investigated the potential roles of StTOC1 in linking environmental signalling and potato tuberisation. We show that transgenic lines with decreased expression of StTOC1 exhibit enhanced StSP6A transcript levels in tuberising stolons, and show changes in gene expression consistent with elevated tuber sink strength.
Project description:Recent advances have defined some of the components of photoperiodic signalling that lead to tuberisation in potato including orthologues of FLOWERING LOCUS T (StSP6A) and CYCLING DOF FACTOR (StCDF1). The aim of the current study is to investigate the molecular basis of permissive tuber initiation under long days in Solanum tuberosum Neo-Tuberosum by comparative analysis with an obligate short day Solanum tuberosum ssp. Andigena accession. We show that the Neo-Tuberosum accession, but not the Andigena, contains alleles that encode StCDF1 proteins modified in the C-terminal region, likely to evade long day inhibition of StSP6A expression. We also identify an allele of StSP6A from the Neo-Tuberosum accession, absent in the Andigena, which is expressed under long days. Other leaf transcripts and metabolites that show different abundances in tuberising and non-tuberising samples were identified adding detail to tuberisation-associated processes. Overall, the data presented in this study highlight the subtle interplay between components of the clock-CONSTANS-StSP6A axis which collectively may interact to fine-tune the timing of tuberisation.
Project description:Heat stress is one of the major abiotic stress factor that affects wheat yield. Especially, heat stress during grain filling affects grain yield besides reduced grain quality. So, in our present study, three genotypes with varied levels of tolerance to heat stress were chosen. They were subjected to heat stress at two stages for three days viz., early (11-14days-post-anthesis) and late (27-30dpa) grain filling independently under controlled conditions. At 14 and 30dpa, the spikes were harvested from control and stress conditions from all the three genotypes, grains were isolated and pulverized. Hence pulverized tissues are used for RNA extraction and further for transcriptome sequencing using HiSeq 4000. Data were analyzed to identify the genes involved in imparting heat stress tolerance.
Project description:The protein content determines the cell state. The variation in protein abundance is crucial when organisms are in the early stages of heat stress, but the reasons affecting their changes are largely unknown. We quantified 47,535 mRNAs and 3,742 proteins in filling grain of wheat under two thermal environments. The impact of mRNA abundance and sequence features which implicated in protein translation and degradation on protein expression was evaluated by regression analysis. Transcription, codon usage and amino acid frequency mainly drive the changes in protein expression under heat stress, and their combined contribution explains 58.2% and 66.4% of protein variation in 30 and 40 °C, respectively. Of which, transcription contributes more to the alteration in protein content under 40 °C (31%) than to 30 °C (6%). Codon usage plays a stable and powerful role in protein expression under heat stress, even surpassing transcription. What’s more, the usage of AAG is a key factor regulating rapid protein expression under heat stress.
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. Experiment 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:Fusarium head blight (FHB) is a major disease of cereal crops caused by the fungus Fusarium graminearum (Fg). FHB affecting the flowering heads (or spikes). A FHB resistance locus has been identified on the chromosome 7E of the wild wheat relative Thinopyrum elongatum (Th.e.). That chromosome (7E) or a long arm fragment of it (7EL) have been transferred as additions in the wheat background 'Chinese Spring' (CS). The two addition lines are resistant to FHB while 'Chinese Spring' is moderately susceptible to it. The mechanism of resistance is not known. The analysis of this work is published in the Canadian Journal of Plant Pathology (Wang et al, 2010). We used the wheat microarray to determine the global expression profil in inoculated spikelets of the addition and parental lines, after water or Fg treatment, with samplings at 2 and 4 days after inoculation (DAI).
Project description:Identify the genetic loci associated with ergot resistance in the cv. Greenshank using a series of transcriptome libraries created from infected doubled haploid individuals segregating for resistance/susceptibility.