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| Author(s): |
Luís Antonio Alves de Toledo Filho
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | Ribeirão Preto. |
| Institution: | Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC) |
| Defense date: | 2012-08-03 |
| Examining board members: |
Maria Helena de Souza Goldman;
Helaine Carrer;
Lazaro Eustaquio Pereira Peres
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| Advisor: | Maria Helena de Souza Goldman |
| Abstract | |
Plant reproduction is a topic of great interest for both basic science and agriculture. The pistil, the angiosperms female reproductive structure, is an organ that plays important roles on the successful reproduction. It is responsible for receiving pollen grains, providing conditions for their hydration and the emission of pollen tubes that grow towards the ovary. In addition, the pistil produces fruits and seeds after the fecundation of the ovules, contained in the ovary. Since a very long time the phytohormone auxin is studied and much is known about its physiological and molecular modes of action. Nevertheless, even though recent advances suggest a great influence of this hormone on the control of the pistil development, there are scarce data in the literature about the influence of elevated levels of auxin on the development of the upper parts of the pistil: the stigma and style. To fill this gap, this project aimed: I) to produce transgenic Nicotiana tabacum plants with elevated auxin levels within the pistil through the expression of the auxin biosynthesis enzyme tryptophan-monoxygenase (iaaM), from Agrobacterium tumefaciens, under the control of the N tabacum STIG1 promoter, which confers stigma-specific expression; II) to analyze the pistil phenotype of the transgenic plants obtained, comparing with wild-type plants, mainly concerning the pistil morphology and reproductive parameters; and III) to characterize through quantitative RT-PCR the expression patterns of different putative auxin-responsive genes in the pistils of transgenic and control plants. A total of 19 stable transgenic plants were obtained, from which 13 presented evident morphological alterations on the pistil. The styles of the STIG1prom::iaaM transgenic plants were longer at the early floral developmental stages, but shorter and thicker at the final stages. Together, these facts suggest a differential mode of perception of elevated auxin levels by the pistil along its development. At anthesis, it was observed a decrease on the stigmatic surface area, with concomitant reduction in the exudate secretion in these plants. Additionally, STIG1prom::iaaM plants presented difficulties in the self-pollination process, and fruits produced by hand-controlled-pollination of transgenic pistils had a reduced final size, making evident the deleterious effects of elevated auxin levels on pistil development and/or on the pollination process. Auxin measurements via LC-MS confirmed a considerable increment in the endogenous levels of this hormone in the pistils of one transgenic plant (STIG1prom::iaaM 4), for the three developmental stages analyzed. Gene expression analyses for 9 putative auxin-responsive genes through qRT-PCR, in pistils from three transgenic plants of three distinct floral developmental stages, revealed significant alterations on the pistil transcriptome of the transgenic plants, suggesting a molecular contribution for some of the analyzed genes on the establishment of the observed phenotypes. Besides providing relevant information concerning the role of auxin in pistil development, the transgenic plants STIG1prom::iaaM, obtained in this project, consist excellent tools for future investigations about the effects of auxin in the regulation of gene expression at the transcriptional and post-transcriptional levels of pistil expressed genes. (AU) | |
| FAPESP's process: | 10/04887-1 - Expression of the Agrobacterium tumefaciens iaaM gene in the Nicotiana tabacum stigma: auxin effects on pistil development |
| Grantee: | Luís Antônio Alves de Toledo Filho |
| Support Opportunities: | Scholarships in Brazil - Master |