Project description:To evaluate the role of seeds in fruit quality, we induced parthenocarpy in tomato by regulating ovule-specific auxin synthesis or responsiveness using the INO promoter from A. thaliana, which is expressed in the outer layer of the integuments during early stages of ovule development. We compared these to fruit where the same coding regions were expressed from the DeFH9 promoter which is expressed in carpel tissues during early stages of ovule development. Expression of auxin synthesis or responsiveness genes by both of these promoters produced seedless parthenocarpic tomato fruit. We compared fruit samples using the Affymetrix tomato GeneChip (GPL4741) to determine how gene regulation and expression differed between wild-type and transgenic fruit. Keywords: genetic modification
Project description:To evaluate the role of seeds in fruit quality, we induced parthenocarpy in tomato by regulating ovule-specific auxin synthesis or responsiveness using the INO promoter from A. thaliana, which is expressed in the outer layer of the integuments during early stages of ovule development. We compared these to fruit where the same coding regions were expressed from the DeFH9 promoter which is expressed in carpel tissues during early stages of ovule development. Expression of auxin synthesis or responsiveness genes by both of these promoters produced seedless parthenocarpic tomato fruit. We compared fruit samples using the Affymetrix tomato GeneChip (GPL4741) to determine how gene regulation and expression differed between wild-type and transgenic fruit. Experiment Overall Design: Wild-type fruit with seeds was compared with transgenic lines INO-IaaM, DefH9-IaaM, INO-RolB, and DefH9-RolB. To find genes with seed-specific expression, we also compared the control with wild-type fruit from which seeds had been manually removed. We had three biological replicates for each treatment and control except DefH9-RolB, for which only two replicates were available. Each CEL file from the microarray represents one plant from each line.
Project description:Early fruit development is crucial for crop production in tomato. After fertilization, the ovary undergoes a cell division and a cell expansion stages before maturation. Although the roles of regulatory signals such as hormone and carbohydrate during early fruit development have been studied, the spatial distribution and the sequential initiation of these regulatory signals is still poorly understood. Using the cultivated tomato “Moneymaker” as materials, we analyzed the transcriptome of the ovule and the ovary wall dissected from the different stages of the early developing fruits. These datasets provide us the whole picture about the spatial and temporal signal distribution of fruit formation which has not been studied in such a detailed manner. Our results suggest that the hormone signal was initiated in both ovule and ovary wall after fertilization. After that, the different signals were activated in ovule and ovary wall due to their distinct developmental processes. By analyzing the global expression profiling of hormone related genes, we found that the auxin might be synthesized in both ovule and ovary wall after fertilization. The expression pattern of sugar related genes revealed the different carbohydrate metabolism events occurred in ovule and ovary wall. At last, we identified a gene showed tissue and stage specific expression pattern and localized in previously reported fruit weight locus which might be selected during tomato breeding.
Project description:Early fruit development is crucial for crop production in tomato. After fertilization, the ovary undergoes a cell division and a cell expansion stages before maturation. Although the roles of regulatory signals such as hormone and carbohydrate during early fruit development have been studied, the spatial distribution and the sequential initiation of these regulatory signals is still poorly understood. Using the cultivated tomato “Moneymaker” as materials, we analyzed the transcriptome of the ovule and the ovary wall dissected from the different stages of the early developing fruits. These datasets provide us the whole picture about the spatial and temporal signal distribution of fruit formation which has not been studied in such a detailed manner. Our results suggest that the hormone signal was initiated in both ovule and ovary wall after fertilization. After that, the different signals were activated in ovule and ovary wall due to their distinct developmental processes. By analyzing the global expression profiling of hormone related genes, we found that the auxin might be synthesized in both ovule and ovary wall after fertilization. The expression pattern of sugar related genes revealed the different carbohydrate metabolism events occurred in ovule and ovary wall. At last, we identified a gene showed tissue and stage specific expression pattern and localized in previously reported fruit weight locus which might be selected during tomato breeding.
Project description:A transcriptome analysis was applied on two peach (Prunus persica L.) cultivars with different sensitivity to low temperature regimes to identify cold-responsive genes that might be involved in tolerance to long low temperature storage. Peach fruit from ‘Morettini No2’ and ‘Royal Glory’, a sensitive and a tolerant, to chilling injury cultivars, respectively, were harvested at commercial maturity stage and allowed to ripen at room temperature (25°C) or subjected to 4 and 6-weeks of cold storage (0°C, 95% R.H.) followed by ripening at room temperature. Microarray experiments, employing the peach microarray platform (μ PEACH 1.0), were carried out by comparing harvested fruit against 4- and 6-week cold-stored fruit. The analysis identified 173 and 313 genes that were differentially expressed in ‘Morettini No2’ and ‘Royal Glory’ fruit after 4 weeks, respectively. However, the 6 weeks cold storage provoked a decrease in the total number of genes differentially expressed in both cultivars. RNA blot analysis validated the differential expression of certain genes showed in microarray data. Among these genes, two heat shock proteins (hsps), a putative β-D-xylosidase, an expansin, a dehydrin and a pathogenesis-related protein PR-4B precursor were induced during cold storage in both cultivars. The induction of hsps and the putative β-D-xylosidase appeared to be independent on the duration of postharvest treatment. On the other hand, transcript levels of lipoxygenase were quite constant during postharvest ripening, while a strong reduction or disappearance was observed after cold storage. A dehydration-induced RD22-like protein showed a reduction in the accumulation of transcripts during postharvest ripening independently on the temperature conditions. Overall, the current study shed some light on the molecular aspects of cold stress in peach fruit quality and identified some ripening and/or cold-induced genes which function need further elucidation.
Project description:‘Crumbly’ fruit is a developmental disorder in red raspberry (Rubus idaeus) that results in malformed fruit with poor adherence of drupelets to one another. In terms of quality and yield, crumbly fruit has become a serious problem in the raspberry industry resulting in unsaleable fruit and waste. A microarray experiment, using pools of progeny from a segregating mapping population (Glen Moy x Latham) with either 'normal' or 'crumbly' fruit at three different fruit developmental stages ('closed'; 'open'; 'green'), identified several genes that were differentially expressed between the crumbly and non-crumbly phenotypes within three quantitative trait loci (QTL) identified. Analysis of gene function highlighted the importance of processes that compromise ovule fertilization as triggers of the crumbly fruit phenotype.
Project description:Fruit set is triggered after ovule fertilization, as a consequence of the downregulation of ovary growth repressors, such as the tomato transcription factors Auxin/indole-3-acetic acid 9 (IAA9) and Agamous-like 6 (AGL6). We produced small RNA libraries from IAA9- and AGL6-silenced ovaries to identify miRNAs differentially expressed in IAA9- and AGL6-silenced ovaries as compared with unpollinated control ovaries. The identified miRNAs represent a pool of regulatory sRNAs potentially involved in tomato fruit initiation.
Project description:As time- and ensemble-averaged measures, NMR observables contain information about both protein structure and dynamics. This work represents a computational study to extract such information for membrane proteins from orientation-dependent NMR observables: solid-state NMR chemical shift anisotropy and dipolar coupling, and solution NMR residual dipolar coupling. We have performed NMR-restrained molecular dynamics simulations to refine the structure of the membrane-bound form of Pf1 coat protein in explicit lipid bilayers using the recently measured chemical shift anisotropy, dipolar coupling, and residual dipolar coupling data. From the simulations, we have characterized detailed protein-lipid interactions and explored the dynamics. All simulations are stable and the NMR restraints are well satisfied. The C-terminal transmembrane (TM) domain of Pf1 finds its optimal position in the membrane quickly (within 6 ns), illustrating efficient solvation of TM domains in explicit bilayer environments. Such rapid convergence also leads to well-converged interaction patterns between the TM helix and the membrane, which clearly show the interactions of interfacial membrane-anchoring residues with the lipids. For the N-terminal periplasmic helix of Pf1, we identify a stable, albeit dynamic, helix orientation parallel to the membrane surface that satisfies the amphiphatic nature of the helix in an explicit lipid bilayer. Such detailed information cannot be obtained solely from NMR observables. Therefore, the present simulations illustrate the usefulness of NMR-restrained MD refinement of membrane protein structure in explicit membranes.
Project description:Flowers have a species-specific fertile period during which pollination and fertilization have to occur to initiate seed and fruit development. Within the flower, the functional life span of the ovule containing the female gametophyte is decisive for fertilization and the initiation of seed development. Here we performed an RNA-sequencing based transcriptome analysis of senescing unfertilized ovules during in a time series. We isolated ovules from Arabidopsis thaliana flowers emasculated at stage 12c at three different time points: 2 days after emasculation (DAE), 3 DAE, and 4 DAE. These time points correspond to intact mature ovules (2DAE), early ovule senescence (3 DAE), and late ovule senescence (4 DAE). We extracted total RNA from the ovules in 3 independent biological replicates, thus generating 9 RNA samples in total, for RNA-sequencing by Illumina HiSeq.
Project description:To distinguish PsCA-responsive proteins, total proteins extracted from the non-acclimated fruit, the fruit exposed to 3 d of cold acclimation, the control fruit stored at 5oC for 12 d and the fruit exposed to 3d of PsCA treatment followed by 9 d of cold storage were separated by 2-DE, respectively. More than 750 protein spots were detected using PDQuest Advanced 2-D Gel Analysis software (Version 8.0) following the 2-DE gel analysis. Of the 750 spots, 58 that displayed significant and reproducible changes in abundance were isolated, and 56 spots were successfully characterized using MS/MS coupled with database searching, which matched 51 proteins. 38 proteins have scores greater than threshold scores of 61(P < 0.05) and they are regarded as the significant differentially- accumulated proteins (SDAPs).