Publicação
RNA-seq analysis of the Quercus suber root response to drought
| Resumo: | Cork oak (Quercus suber L.) is a typical species from the Portuguese Mediterranean forest, usually growing in dry environments. Mainly due to its high economic value, cork oak has been considered a protected plant species and a national species of interest. Factors resulting from climate changes, e.g. increased temperature, high light intensity, drought and air pollution are directly affecting the development of plants and reducing their productivity. Next generation sequencing, in particular RNAseq, has proven to be a powerful tool with a diverse range of applications, and provides fundamental insight into the structure and regulatory aspects of gene networks, particularly when organisms without avalible OMICs resources, such as a sequence genome, are studied. To address the cork oak’s response and regulation to water availability, differential transcriptomic analysis was performed in roots of young plantlets subjected to moderate and high drought conditions, comparing to well-irrigated plants. Photosynthetic activity measured by PAM fluorometry and photosynthetic pigment content were used as physiological indicators of plant fitness. cDNA was obtained from previously droughtstressed roots RNA, and sequencing was carried out by Roche’s 454 technology resulting in three libraries, of a 1.8 million reads in total, later assembled into 21012 unigenes, that would be used for mapping steps, in order to identify differential expression. This resulted in the identification of 353 up-regulated and 193 down-regulated genes. Primary in silico analysis identified a significant number of effector genes traditionally associated with drought responses, such as dehydrins, LEA proteins, cell wall remodeling, transcription factors and ubiquiting-associated genes, suggesting a tight control of drought responses at both the transcriptional and protein turnover levels. Further in silico analysis was accomplished by annotating the genes against the genome of the model plant Arabidopsis thaliana, allowing for ortholog identification. This allowed us to establish gene networks based on functional characteristics, such as gene co-expression, protein-protein interactions and co-localization. Coupled with ciselement enrichment and transcription factor identification, this analysis enabled the establishment of functional and regulatory relationships between differentially expressed genes. Ultimately, we demonstrated the induction, in drought stressed Quercus suber roots, of a complete, ABA-dependent signaling cascade. Elements of this signaling cascade range from ABA-sensing components to transcription factors and then to effector genes involved in the drought response. |
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| Autores principais: | Magalhães, Alexandre Miguel Papadopoulos |
| Assunto: | Ciências Naturais::Ciências Biológicas |
| Ano: | 2015 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | Cork oak (Quercus suber L.) is a typical species from the Portuguese Mediterranean forest, usually growing in dry environments. Mainly due to its high economic value, cork oak has been considered a protected plant species and a national species of interest. Factors resulting from climate changes, e.g. increased temperature, high light intensity, drought and air pollution are directly affecting the development of plants and reducing their productivity. Next generation sequencing, in particular RNAseq, has proven to be a powerful tool with a diverse range of applications, and provides fundamental insight into the structure and regulatory aspects of gene networks, particularly when organisms without avalible OMICs resources, such as a sequence genome, are studied. To address the cork oak’s response and regulation to water availability, differential transcriptomic analysis was performed in roots of young plantlets subjected to moderate and high drought conditions, comparing to well-irrigated plants. Photosynthetic activity measured by PAM fluorometry and photosynthetic pigment content were used as physiological indicators of plant fitness. cDNA was obtained from previously droughtstressed roots RNA, and sequencing was carried out by Roche’s 454 technology resulting in three libraries, of a 1.8 million reads in total, later assembled into 21012 unigenes, that would be used for mapping steps, in order to identify differential expression. This resulted in the identification of 353 up-regulated and 193 down-regulated genes. Primary in silico analysis identified a significant number of effector genes traditionally associated with drought responses, such as dehydrins, LEA proteins, cell wall remodeling, transcription factors and ubiquiting-associated genes, suggesting a tight control of drought responses at both the transcriptional and protein turnover levels. Further in silico analysis was accomplished by annotating the genes against the genome of the model plant Arabidopsis thaliana, allowing for ortholog identification. This allowed us to establish gene networks based on functional characteristics, such as gene co-expression, protein-protein interactions and co-localization. Coupled with ciselement enrichment and transcription factor identification, this analysis enabled the establishment of functional and regulatory relationships between differentially expressed genes. Ultimately, we demonstrated the induction, in drought stressed Quercus suber roots, of a complete, ABA-dependent signaling cascade. Elements of this signaling cascade range from ABA-sensing components to transcription factors and then to effector genes involved in the drought response. |
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