Valentine, Tracey Anne; (1999) Isolation of Arabidopsis mutants resistant to root pattern disrupting signals from carrot embryogenic cultures. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

During development, individual plant cells differentiate into specific cell types, creating characteristic cellular differentiation patterns. Plants use both positional information and cell-cell signalling in constructing these patterns. The nature of the cell-cell signals are unknown.Cell to cell signalling is involved in the early stages of somatic embryogenesis with arabinogalactan-proteins (AGPs) being essential to the process. The type of AGPs expressed at the cell wall of cells undergoing embryogenesis are linked to the embryogenic potential of the cell and also the cell-cell signal secreted by a cell. AGPs.are found exposed at the plasma membrane, in intravacuolar vesicles and in the extracellular matrix of plant cells. Monoclonal antibodies have been used to designate different type of AGPs. Using these, several AGPs have been found to be expressed in both embryogenic cultures and in the plant embryos and roots, with the expression patterns of some AGPs relating to, or predict cellular differentiation patterns. The signals involved in the embryogenic process can be obtained from the culture media or by washing the cell walls with water. Extracts from somatic embryogenic cultures (CWSE) containing AGP like carbohydrates were shown to affect the root development of both Carrot and Arabidopsis seedlings. In response to CWSE, seedlings produced roots, which were shorter and fatter than controls with lateral root production also affected. There was an increase, in the number of cell layers, and in the number of cells in each cell layer of the root. Putative signals have been purified using a combination of the effect of the CWSE on Arabidopsis roots and separation columns. Purification was achieved by fractionation using a Sephadex G-10 column followed by molecular weight cut off membranes and partition chromatography using a Flash Biotage unit. 5 active fractions have been analysed for carbohydrate composition. Several unusual residues were found, including inositol 0-methyl ethers. The similarity of the residues found in each of the fractions, raises the possibility that the compounds are part of a biochemical pathway and that some of the smaller molecules may be processed sections of the larger molecules. Several putative mutants that had reduced sensitivity to the CWSE, were isolated from M2 populations oiArabidopsis derived from ethane methane sulphanate treated plants. The seedlines were retested for their resistance to CWSE against their control grown phenotype. The mutants isolated tended to have shorter roots than control grown coll, gll plants, and produce a reduced number of lateral roots.

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