Identificação de fatores de transcrição que controlam a expressão do gene GmASR3 de soja

• Autor(a) principal: Dadalto, Silvana Pinheiro
• Data de Publicação: 2012
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/2453
• topic: GmASR3; Soja; Proteína-DNA; GmASR3; Soybean; Protein-DNA; CNPQ::CIENCIAS BIOLOGICAS::BIOQUIMICA::BIOLOGIA MOLECULAR

ASR Proteins are ABA, stress and ripening induced proteins present in various plant species such as lily (Lilium longiflorum), sugarcane (Saccharum officinarum), grape (Vitis vinifera) ; Rice (Oryza sativa), petunia, banana (Musa acuminata and Musa balbisiana) and corn (Zea mays). However no single ASR gene has been found in Arabidopsis. ASRs are small, hydrophilic and unstructured proteins. Its function, as well as the signaling pathway to which they belong, remains unknown. In soybean were reported the presence of three ASRs, which were called GmASR1, and GmASR2 GmASR3. This study aimed to identify factors that interact with the GmASR3 promoter region, as well as factors that bind to the protein GmASR3. After analysis of databases, the promoter region about 500 bp upstream of the transcription initiation codon was obtained from PCR performed with genomic DNA from soybean (Glycine max), and cloned into plasmid pHIS2.1. This construct was transformed in yeast S. cerevisiae W303 for subsequent liquid screening of cDNAs that express factors capable of interacting with the promoter GmASR3 by mono-hybrid yeast system. It was found that the protein GmASR3 has weak transactivation capacity under the used conditions, which enabled the realization of two-hybrid experiments to identify factors that bind to the protein GmASR3.The factors that bind to GmASR3 were obtained by two-hybrid system yeast, using for this yeast S. cerevisiae AH109 carrying the plasmid pDest32 which was inserted the cDNA sequence GmASR3. Likewise, the method of liquid screening in deep well plates was used. The colonies that were considered to have the factors that interact with the promoter or the protein in GmASR were re-transformed in E. coli Dh5α and sent for sequencing.