Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of trans-differentiation of cultured Arabidopsis cells

ABSTRACT: The formation of vascular tissue occurs when cellulose, hemicellulose, lignin and other wall components are deposited within the primary cell wall. These secondary thickened cells then undergo programmed cell death producing a network of empty cells with which water and ions can be transported throughout the plant. The hormones auxin and cytokinin are the principle signals for vascular tissue initiation. As a consequence cells cultured in-vitro can be converted into vascular tissue with the addition of exogenous auxin and cytokinin. We have created an in-vitro cell system, using callus produced from leaves that can be induced to form vascular tissue. Leaves are callused on induction media for two weeks. The callus is then transferred to liquid media and incubated under optimum conditions resulting in an increase in vascular tissue formation. Approximately 20% of cells will differentiate during the incubation period. The alteration of cytokinin concentration affects the ability of the cultured cells to undergo differentiation. Consequently callus incubated in liquid media, containing lower cytokinin concentrations, will undertake relatively little differentiation. Samples have been isolated from cell cultures at different time points and different hormone concentrations during incubation. Quantitative PCR using the marker AtCesA7, which encodes a cellulose synthase subunit specific to secondary wall deposition, was used as a guide to determine periods of high and low vascular differentiation. This system provides an opportunity to compare gene expression between differentiating and non differentiating cells and allow the identification of genes up regulated during vascular tissue formation.

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: unknown unknown

PROVIDER: E-NASC-15 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Agrobacterium tumefaciens, a bacterial species found in temperate soils world wide, is the causative agent of crown gall disease on many plants. A. tumefaciens-induced tumours are feared in orchards and vineyards because of their pathological interference with nutrient and water supply which results in crop decline. Small wounds at the crown of the plant, usually induced by wind-bending, are potential entry sites for the bacterium. The tumorous growth is initiated by the integration and expression of the T-DNA of the bacterial Ti plasmid within the plant nuclear DNA. The T-DNA encodes enzymes catalysing the synthesis of increased concentrations of auxin and cytokinin, and of opines which stimulate cell division and enlargement. The fast growing tumours have been shown to be a strong nutrient sink on their host plants. As a matter of fact, sugar and K+ content were found to be up to 10- and 5-fold, respectively, higher in this tissue and transpiration was about 15 times increased compared to normal tissue. Whereas the morphological structure as well as some physiological and biochemical parameters of the tumour have been analysed in detail, little is known about the underlying gene expression pattern. Proliferation and growth of the tumour induced by Agrobacterium tumefaciens is obviously due to the extraordinary high concentration of phytohormons, minerals and metabolites. Their influence on regulation of gene transcription will provide information on the mechanisms underlying fast tumour growth. In a project funded by the DFG we recently started to investigate the role of solute transporter for tumour development on the model plant Arabidopsis thaliana. By comparing the expression pattern of RNA preparations from Arabidopsis tumour and non-tumour tissue, we will be able to identify genes which facilitate crown gall development. For the expression analysis with an Affymetrix full genome chip we will induce tumours at the base of an injured Arabidopsis inflorescence stalk (var. Wassilewskija, WS-2). RNA will be extraxted from tumour and injured non-tumor inflorescence stalk tissue using the RNeasy Plant Mini Kit (Qiagen), followed by a DNase treatment to eliminate DNA contamination.

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Transcription profiling of trans-differentiation of cultured Arabidopsis cells

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