Project description:Sunflower is an important source of vegetable oil worlwide. A differential organ-specific sunflower ESTs was previously generated by a subtractive hybridization method, including a considerable number of abiotic stress associated sequences. The objective of this work is to analyze the sunflower gene expression of previously identified candidate genes under a comprehensive microarray analysis of the leaf transcriptoma under cold and salinity stresses, considering the impacts of these abiotic stresses on sunflower yield in many productive areas. The aimed of this work is to perform genome analysis of sunflower based on its functional regions and the characterization of the sunflower transcriptoma profiles for different organ-specific genes. Abiotic-related expressed genes were the target of this characterization through a gene expression analysis of the local EST bank (annotated according to Gene Ontology Annotation) using a cDNA organ-specific microarray chip approach. We analyzed 287 differentially expressed genes derived from leaf, stem, R1 and R4 flower developmental stages. Transcriptional analysis allowed the detection of three different groups of genes according to their expression patterns. Group 1 contained 112 up-regulated genes under abiotic stress conditions (cold or salinity), whereas Group 2 (126 genes) did not show changes in their expression levels. On the other hand, 49 genes were classified as Group 3 included were down-regulated genes under both stresses. Eighty genes exhibited a significative fold change under abiotic stress conditions, being six of them validated by qRT-PCR. Microrarray profiling of cold and NaCl-treated sunflower leaves revealed dynamic changes in transcript abundance, including transcription factors, defense/stress related proteins, and effectors of homeostasis, all of which highlight the complexity of both stress responses. This finding provides identification of many transcriptional processes occurring under abiotic stress in sunflower for genes isolated from organ-specific cDNA libraries Keywords: gene expression profile of organ-specific sunflower transcriptoma in response to NaCl and cold

Project description:Sunflower is an important source of vegetable oil worlwide. A differential organ-specific sunflower ESTs was previously generated by a subtractive hybridization method, including a considerable number of abiotic stress associated sequences. The objective of this work is to analyze the sunflower gene expression of previously identified candidate genes under a comprehensive microarray analysis of the leaf transcriptoma under cold and salinity stresses, considering the impacts of these abiotic stresses on sunflower yield in many productive areas. The aimed of this work is to perform genome analysis of sunflower based on its functional regions and the characterization of the sunflower transcriptoma profiles for different organ-specific genes. Abiotic-related expressed genes were the target of this characterization through a gene expression analysis of the local EST bank (annotated according to Gene Ontology Annotation) using a cDNA organ-specific microarray chip approach. We analyzed 287 differentially expressed genes derived from leaf, stem, R1 and R4 flower developmental stages. Transcriptional analysis allowed the detection of three different groups of genes according to their expression patterns. Group 1 contained 112 up-regulated genes under abiotic stress conditions (cold or salinity), whereas Group 2 (126 genes) did not show changes in their expression levels. On the other hand, 49 genes were classified as Group 3 included were down-regulated genes under both stresses. Eighty genes exhibited a significative fold change under abiotic stress conditions, being six of them validated by qRT-PCR. Microrarray profiling of cold and NaCl-treated sunflower leaves revealed dynamic changes in transcript abundance, including transcription factors, defense/stress related proteins, and effectors of homeostasis, all of which highlight the complexity of both stress responses. This finding provides identification of many transcriptional processes occurring under abiotic stress in sunflower for genes isolated from organ-specific cDNA libraries To control biological variation between individuals, three biological samples from the same tissue were pooled on one sample prior to probe preparation. The reference (control) sample consisted of pooled RNA extracted from sunflower seedlings growing under unaltered environmental greenhouse conditions, whereas chilling and salinity samples were RNA extracted from sunflower seedlings growing in greenhouse under those stressed conditions. The RNA (800ng) samples were labeled by using SuperScript Indirect RNA Amplification System Kit (Invitrogen, cat# L1016-02) based on the method designed by Eberwine y col. 1992. Following RNA amplification (with the incorporation of UTP aminoallil), labeled product was achieved by incubating with Cy3 or Cy5 esters in alkaline media. Slides were used in order to quantify the relative expression of ESTs in control and treated leaves by Cy3 and Cy5 hybridization technique Dye-swaps were used to correct for differences in incorporation and fluorescent properties of both dyes, generating a number of 9 slides per experiment (three slides for control and three slides for each treatment) with a total number of 18 slides considering technical replicates.

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