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| Author(s): |
Irving Joseph Berger
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Piracicaba. |
| Institution: | Universidade de São Paulo (USP). Escola Superior de Agricultura Luiz de Queiroz (ESALA/BC) |
| Defense date: | 2005-05-23 |
| Advisor: | Ricardo Antunes de Azevedo |
| Field of knowledge: | Biological Sciences - Biochemistry |
| Indexed in: | Banco de Dados Bibliográficos da USP-DEDALUS |
| Location: | Universidade de São Paulo (USP). Escola Superior de Agricultura Luiz de Queiroz; ESALQ-BC /t635.642; B496t |
| Abstract | |
Tomato (Lycopersicon esculentum Mild.) chloroplast transformation in 2001, more than one decade after the first results from the transplastomic technology in tobacco, has increased the opportunity to the introduction and expression of transgenes into plastid genomes of crop species. Among other advantages, environmental safety and high levels of transgene expression with consequent great accumulation of the introduced recombinant protein are results of the plastid transformation. Plastids take part in several metabolic cellular processes and so modification of any of the involved pathways could be viable by the introduction of exogenous genes or alteration of endogenous genes in chloroplast genomes. Since vitamin E, specially α-tocopherol, has important therapeutic properties when daily intakes exceed normal nutritional requirements (100 to 400 IU), the goal of this work was the production of vitamin E rich plants by chloroplast transformation. ln that way, the Synechocystis sp. PCC6803 s/r0089 and s/r0090 genes coding for the y-tocopherol methyl transferase (γ-TMT) and HPP dioxygenase (HPPDase) enzymes respectively were isolated and introduced in chloroplast transformation vectors resulting in the plJB30, plJB31 and plJB32 plasmids, which contain the s/r,0089, s/r,0090 and s/r0089+s/r0090 coding regions respectively. Biolistic experiments with tomato and tobacco leaf explants were performed for the integration and expression of the constructs into the plastid genomes. Transformed tobacco plants containing all the three constructs were confirmed to be homoplasmic and have the transgenes correctly integrated. Since overexpression of the HPPDase coding genes in plants can lead to resistance against HPPD inhibiting herbicides, transplatomic tobacco plants were analized and have shown resistance against isoxaflutole on in vitro tests. Tomato homoplasmic plants containing the transgenes were confirmed only for the plJB32 construct and have also the transgenes correctly integrated. (AU) | |