Arabidopsis thaliana
Ontology highlight
ABSTRACT:
OTHER RELATED OMICS DATASETS IN: E-GEOD-13783
PROVIDER: PRJNA110613 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Transient genetic modification of plant protoplasts is a straightforward and rapid technique for the analysis of numerous aspects of plant biology. One drawback in the analysis of transformed protoplast suspensions is that they are a heterogeneous mix of cells that have and have not been successfully transfected. To overcome this problem, we have developed a system that employs a fluorescent positive selection marker in combination with flow cytometric analysis as well as fluorescence activated cell sorting (FACS) to isolate responses in the transfected protoplasts exclusively. This recombinase-compatible system enables high-throughput screening of genetic circuitry. Moreover, the use of FACS allows in depth downstream analysis. Lastly, over-expression is an effective means to dissect regulatory networks, especially where redundancy exists. Here, this system has been applied to the study of auxin signaling in order to investigate reporter gene activation and genome-wide transcriptional changes in response to manipulation of the auxin-response network. We have transiently over-expressed dominant negative mutant isoforms of Aux/IAA transcription factors (IAA7mII and IAA19mII; Tiwari et al., 2001) in Arabidopsis Pwer::GFP root protoplasts, making use of a RFP fluorescent positive selection marker and FACS to isolate the dually labeled (IAAnmII expressing and Pwer::GFP-positive) cells. We have compared the transcriptional differences between an empty vector control, IAA7mII and IAA19mII protoplasts that had either been treated with 5microM IAA or mock-treated for 3 hours. Keywords: hormone treatment, genetic modification
2009-03-01 | GSE13783 | GEO
Project description:Transient genetic modification of plant protoplasts is a straightforward and rapid technique for the analysis of numerous aspects of plant biology. One drawback in the analysis of transformed protoplast suspensions is that they are a heterogeneous mix of cells that have and have not been successfully transfected. To overcome this problem, we have developed a system that employs a fluorescent positive selection marker in combination with flow cytometric analysis as well as fluorescence activated cell sorting (FACS) to isolate responses in the transfected protoplasts exclusively. This recombinase-compatible system enables high-throughput screening of genetic circuitry. Moreover, the use of FACS allows in depth downstream analysis. Lastly, over-expression is an effective means to dissect regulatory networks, especially where redundancy exists. Here, this system has been applied to the study of auxin signaling in order to investigate reporter gene activation and genome-wide transcriptional changes in response to manipulation of the auxin-response network. We have transiently over-expressed dominant negative mutant isoforms of Aux/IAA transcription factors (IAA7mII and IAA19mII; Tiwari et al., 2001) in Arabidopsis Pwer::GFP root protoplasts, making use of a RFP fluorescent positive selection marker and FACS to isolate the dually labeled (IAAnmII expressing and Pwer::GFP-positive) cells. We have compared the transcriptional differences between an empty vector control, IAA7mII and IAA19mII protoplasts that had either been treated with 5microM IAA or mock-treated for 3 hours. Experiment Overall Design: 18 samples with 3 replicates for each condition and transformation vector: 3x empty vector mock treated, 3x empty vector IAA treated, 3x IAA7mII over-expressor mock treated, 3x IAA7mII over-expressor IAA treated, 3x IAA19mII over-expressor mock treated and 3x IAA19mII over-expressor IAA treated.
2009-03-07 | E-GEOD-13783 | biostudies-arrayexpress
Project description:This work studies the impact of AtNIGT1/HRS1-GR entrance in the nucleus upond DEX treatment in protoplasts. AtNIGT1/HRS1 TARGET. The whole procedure has been performed as previously described in Bargmann et al Mol Plant 2013. In brief, the protoplasts were transfected with the plasmid pBeaconRFP_GR-HRS1 that trigger the expression of HRS1 protein fused with the glucocorticoid receptor under control of CaMV35S promoter. Protoplasts were treated with 35µM cycloheximide (CHX) to inhibit translation and to select only direct target genes and 10µM dexamethasone (DEX) to induce HRS1-GR entry in the nucleus. Nitrate is maintained during the whole TARGET procedure. The Red Fluorescent Protein was used as marker selection for fluorescent-activated cell sorting (FACS) of successfully transformed protoplasts. RNA were extracted and amplified in order to be tested with ATH1 Affymetrix™ chips.
2015-01-05 | E-GEOD-64352 | biostudies-arrayexpress
Project description:A fluorescent plant pathogen
| PRJNA19911 | ENA
Project description:Multicellular organisms such as plants contain different types of cells with specialized functions. Analyzing the characteristics of each type of cells will reveal specific cell functions and enhance understanding of how an organism organizes and works. Here we report a high-sensitive and efficient cell-type-specific multi-omics pipeline, combining simplified flow cytometry-based fluorescent cell-sorting for fluorescent protoplasts and optimized nano-scale proteomics and metabolomics methods, which allow us to in-depth analyze the proteomics and metabolomics of a particular type of cells. By this method, we quantitatively compared the proteomics and metabolomics between guard cells and mesophyll cells and revealed that the enrichment of signal transduction-related proteins enables guard cells to respond to various environmental stimuli quickly. We uncovered a guard-cell-specific kinase cascade consisting of RAF15 and OST1 mediates the ABA-induced stomatal closure in guard cells. This pipeline is applicable to various types of cells in plant or non-plant systems to acquire systemic knowledge of how cells work specifically and in highly organized multiple cell organisms.
2022-12-18 | PXD030511 | JPOST Repository
Project description:Multicellular organisms such as plants contain different types of cells with specialized functions. Analyzing the characteristics of each type of cells will reveal specific cell functions and enhance understanding of how an organism organizes and works. Here we report a high-sensitive and efficient cell-type-specific multi-omics pipeline, combining simplified flow cytometry-based fluorescent cell-sorting for fluorescent protoplasts and optimized nano-scale proteomics and metabolomics methods, which allow us to in-depth analyze the proteomics and metabolomics of a particular type of cells. By this method, we quantitatively compared the proteomics and metabolomics between guard cells and mesophyll cells and revealed that the enrichment of signal transduction-related proteins enables guard cells to respond to various environmental stimuli quickly. We uncovered a guard-cell-specific kinase cascade consisting of RAF15 and OST1 mediates the ABA-induced stomatal closure in guard cells. This pipeline is applicable to various types of cells in plant or non-plant systems to acquire systemic knowledge of how cells work specifically and in highly organized multiple cell organisms.
2022-12-17 | PXD030489 | JPOST Repository
Project description:T cell development relies on the precise developmental control of various cellular functions for appropriate positive and negative selection. Previously, gene expression profiling of peptide-driven negative selection events in the N15 TCR class I MHC-restricted mouse and D011.10 TCR class II MHC-restricted mouse has offered insights into the coordinate engagement of biological processes affecting thymocyte development. However, there has been little comparable detailed in vivo global genome expression analysis reported for positive selection. We used microarrays to identify the genes differentially expressed during CD8 single positive T cell development in N15 TCR transgenic Rag2 deficient mice.
2009-03-31 | GSE13493 | GEO
Project description:Changes in gene expression form a crucial part of the plant response to pathogen infection. Whole-leaf expression profiling has played a valuable role in identifying genes and processes that contribute to the interactions between the model plant Arabidopsis thaliana and a diverse range of pathogens. However, for highly localised infections, such as downy mildew caused by the biotrophic oomycete pathogen Hyaloperonospora arabidopsidis (Hpa), whole-leaf profiling may fail to capture the complete Arabidopsis response. Highly localised expression changes may be diluted by the comparative abundance of non-responding leaf cells or the Hpa oomycete evading detection by cells. Furthermore, local and systemic Hpa responses of a differing nature may become convoluted. To address this we applied the technique of Fluorescence Activated Cell Sorting (FACS), typically used for analyzing plant abiotic responses, to the study of plant-pathogen interactions. Using the promoter of Downy Mildew Resistant 6 (DMR6) linked to GFP as a fluorescent marker of pathogen-contacting cells, we isolated Hpa-proximal and Hpa-distal cells from infected leaf samples using FACS, and measured global gene expression. The whole experiment was carried out in triplicate, with two time points (5 and 7 days post-inoculation) and three cell types (pathogen-proximal, pathogen-distal and uninfected control), totalling 18 microarrays. Transgenic Arabidopsis thaliana Col-0 with the transgene pDMR6:GFP was used for all experiments. Seedling populations were inoculated with 30,000-60,000 spores/ml of Hyaloperonospora arabidopsis strain Noks1 at 7 days old, and overground tissue sampled at 5 and 7 days post-inoculation. Protoplasts were generated from these samples according to Grønlund et al. 2012 JOVE, and sorted by fluorescence activated cell sorting into a GFP-positive (pathogen-proximal) and GFP-negative (pathogen-distal) population. As a control, uninfected seedlings were also sampled at 12 and 14 days old (equivalent age of 5 and 7 days post-inoculation), protoplasts generated and sorted to yield a GFP-negative population of uninfected control cells. Gene expression was analysed using NimbleGen 12 x 135K microarrays, designed for the TAIR10 genome. Data was normalised using the Robust Multichip Averaging algorithm.
2015-07-13 | E-GEOD-67100 | biostudies-arrayexpress
Project description:Plant cells exhibit remarkable plasticity of their differentiation states, enabling regeneration of whole plants from differentiated somatic cells. How they revert cell fate and express pluripotency, however, remains unclear. In this study we demonstrate that transcriptional activation of auxin biosynthesis is crucial for reprogramming differentiated Arabidopsis leaf cells. We demonstrate that interfering with the activity of histone acetyltransferases dramatically reduces callus formation from leaf mesophyll protoplasts. Histone acetylation permits the transcriptional activation of PLETHORAs (PLTs), leading to the induction of their downstream target gene YUCCA1 (YUC1) encoding an enzyme for auxin biosynthesis. Auxin biosynthesis is in turn required to accomplish initial cell division through the activation of G2/M phase genes mediated by MYB DOMAIN PROTEIN 3-RELATED (MYB3Rs). We further show that the AUXIN RESPONSE FACTOR 7 (ARF7)/ARF19 and INDOLE-3-ACETIC ACID INDUCIBLE 3 (IAA3)/IAA18-mediated auxin signaling pathway is responsible for cell cycle reactivation by transcriptionally upregulating MYB3R4. These findings provide a mechanistic model of how differentiated plant cells revert their fate and reinitiate the cell cycle to become pluripotent.
2022-07-07 | GSE204767 | GEO
Project description:The evolution and diversification of proteins capable of remodelling domains has been critical for transcriptional reprogramming during cell fate determination in multicellular eukaryotes. Chromatin remodelling proteins of the CHD3 family have been shown to have important and antagonistic impacts on seed development in the model plant, Arabidopsis thaliana, yet the basis of this functional divergence remains unknown. In this study, we demonstrate that genes encoding the CHD3 proteins PICKLE (PKL) and PICKLE-RELATED 2 (PKR2) originated from a duplication event during the diversification of crown Brassicaceae, and that these homologues have undergone distinct evolutionary trajectories since this duplication, with PKR2 fast-evolving under positive selection, while PKL is evolving under purifying selection. We find that the rapid evolution of PKR2 under positive selection reduces the encoded protein’s intrinsic disorder, possibly suggesting a tertiary structure configuration which differs from that of PKL. Our whole genome transcriptome analysis of gene expression in seeds of pkr2 and pkl mutants reveals that they act antagonistically on the expression of specific sets of genes, providing a basis for their differing roles in seed development. Our results provide insights on gene duplication and neofunctionalization can lead to differing and antagonistic selective pressures on transcriptomes during plant reproduction, as well as on the evolutionary diversification of the CHD3 family within seed plants.
2023-09-06 | GSE236025 | GEO