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Use of transgenic plants in functional analysis of plant genes Functional genomics makes it possible to detect the functions of uncharacterized genes. It allows you to understand not only the functions of individual genes, but also the relationships between them. The acquired knowledge can also be used in improving the properties of various cultivated plants. With decreasing cost and increasing sequencing efficiency, many plant genomes are sequenced, but without the associated function of individual genes. Reverse genetics approaches are used to characterize these genes. In the first place, these include approaches that include the analysis of insertional mutants or the use of RNA interference that inactivates or reduces the expression of the individual genes studied. We can also inactivate whether to specifically modify the function of a particular selected gene using site-specific endonucleases, which include meganucleases, zinc finger nucleases, TALEN (transcription activator-like effector nucleases) and CRISPR (clustered regularly interspaced short paindromic repeats). In contrast to inactivation approaches, there are approaches such as activating mutagenesis and ectopic expression, through which we increase the expression. In addition, gene expression and protein localization can be analyzed...

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2019-07-18T16:42:05Z No. of bitstreams: 1 2015 - Cassia Pereira Coelho.pdf: 4393249 bytes, checksum: fb85cca04b6e74dd8ba752222181b30d (MD5) Made available in DSpace on 2019-07-18T16:42:05Z (GMT). No. of bitstreams: 1 2015 - Cassia Pereira Coelho.pdf: 4393249 bytes, checksum: fb85cca04b6e74dd8ba752222181b30d (MD5) Previous issue date: 2015-02-12 CAPES FAPERJ CNPq The main goal of the study was to elucidate the functions of the OsAAP1 and OsAAP18 genes and check its influence on the N remobilization to grains filling and storage of proteins in rice plants. To this, were performed experiments to: (1) Phenotypic characterization of T-DNAi nsertional mutant plants of Dong Jin rice varietyand (2) check affinity of transport for amino acids, through functional complementation of yeast mutant strains 22 8AA, unable to transport efficiently L-proline, L-citrulline, L-aspartate and L.glutamate, 22?6AAL, unable to transport efficiently L-lysine and JA248, unable to transport efficiently L-glutamine. The phenotypic analyze of the mutant plants was developed through a greenhouse experiment using theT-DNAs insertonal mutants lines 3A-00581, mutant for the gene LOC_Os07g04180 (OsAAP1), and 4A-00262, mutant for LOC_Os0636210 gene (OsAAP18) and wild type plants.Complementation tests with mutants yeasts were performed on nitrogen-free medium supplemented with L-proline, L-citrulline, L-aspartate, L-glutamate, L-lisyne or L-glutamine as the sole nitrogen source. Frm the results it was concluded that genes OsAAP1 e OsAAP18 encode to proteins involved in nitrogen remobilization to gains in rice plants and OsAAP18 influences rice productivity protein content in grains. O principal objetivo do trabalho foi elucidar a fun??o dos genes OsAAP1 e OsAAP18 e verificar sua influ?ncia sobre a remobiliza??o de N para os gr?os e forma??o de prote?nas de reserva em plantas de arroz. Para isso, foram realizados experimentos para (1) caracteriza??o fenot?pica de plantas de arroz mutantes por inser??o de T-DNA da variedade Dong Jin e (2) verifica??o da afinidade dos transportadores codificados pelos genes OsAAP1 e OsAAP18 por diferentes substratos (amino?cidos) atrav?s de trabalho de complementa??o em leveduras mutantes das cepas 22?8AA, que ? incapaz de transportar eficientemente L-prolina, L-citrulina, L-aspartatoand L-glutamato, 22?6AAL, que ? incapaz de transportar eficientemente L-lisina e JA248, que ? incapaz de transportar eficientemente L-glutamina, carregando a sequ?ncia codantedos genes OsAAP1 e OsAAP18. A an?lise fenot?pica das plantas mutantes foi realizada atrav?s de experimento conduzido em casa de vegeta??o na UFRRJ utilizando-se as linhagens de plantas mutantes por T-DNA para o gene LOC_Os07g04180 (OsAAP1), 3A- 00581, e para o gene LOC_Os0636210 (OsAAP18), 4A-00262 e plantas tipo selvagem como controle. Testes de complementa??o com as leveduras mutantes foram realizados na Washington State University, utilizando-se meio livre de N, suplementado com L-prolina, L-citrullina, L-aspartato, L-glutamato, L-lisina ou L-glutamina como ?nica fonte de N. Ao final do estudo verificou-se que os genes OsAAP1 e OsAAP18 codificam prote?nas de transporte de amino?cidos envolvidas na remobiliza??o de N para os gr?os em plantas de arroz e OsAAP18influ?ncia o metabolismo de plantas de arroz e a produ??o de gr?os.

When plants are exposed to stressful environmental conditions, the production of Reactive Oxygen Species (ROS) increases and can cause significant damage to the cells. Antioxidant defenses, which can detoxify ROS, are present in plants. A major hydrogen peroxide detoxifying system in plant cells is the ascorbate-glutathione cycle, in which, ascorbate peroxidase (APX) enzymes play a key role catalyzing the conversion of H(2)O(2) into H(2)O, using ascorbate as a specific electron donor. Different APX isoforms are present in distinct subcellular compartments, such as chloroplasts, mitochondria, peroxisome, and cytosol. The expression of APX genes is regulated in response to biotic and abiotic stresses as well as during plant development. The APX responses are directly involved in the protection of plant cells against adverse environmental conditions. Furthermore, mutant plants APX genes showed alterations in growth, physiology and antioxidant metabolism revealing those enzymes involvement in the normal plant development.

When plants are exposed to stressful environmental conditions, the production of Reactive Oxygen Species (ROS) increases and can cause significant damage to the cells. Antioxidant defenses, which can detoxify ROS, are present in plants. A major hydrogen peroxide detoxifying system in plant cells is the ascorbate-glutathione cycle, in which, ascorbate peroxidase (APX) enzymes play a key role catalyzing the conversion of H2O2 into H2O, using ascorbate as a specific electron donor. Different APX isoforms are present in distinct subcellular compartments, such as chloroplasts, mitochondria, peroxisome, and cytosol. The expression ofAPX genes is regulated in response to biotic and abiotic stresses as well as during plant development. The APX responses are directly involved in the protection of plant cells against adverse environmental conditions. Furthermore, mutant plants APX genes showed alterations in growth, physiology and antioxidant metabolism revealing those enzymes involvement in the normal plant development.