Unknown,Transcriptomics,Genomics,Proteomics

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Expression profiling of PEG induced water stress responsive genes in Eucalyptus grandis

ABSTRACT: The study was conducted to identify differentially expressed polyethylene glycol (PEG) induced water stress responsive genes in E. grandis. Forty day old rooted cutting of E. grandis was subjected to -0.225 MPa PEG treatment and total RNA was isolated from leaves of water treated control and PEG treated samples after three hours of treatment. The differential expression of water stress responsive genes was analyzed using microarray technique. Agilent two-color experiment, Organism: Eucalyptus,Custom Agilent Eucalyptus 8x60k Microarray Gene expression (AMADID: 59849 ) designed by Genotypic Technology Pvt.Ltd.

ORGANISM(S): Eucalyptus grandis

SUBMITTER: Genotypic technology

PROVIDER: E-GEOD-73106 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Rooting capability is one of the economically traits lost during the juvenile to mature phase change in woody plants. A comprehensive microarray analysis was performed to compare the profiles of gene expression in juvenile and mature cuttings from E. grandis, either auxin treated or untreated on days, 0, 1, 3, 6, 9 and 12 post excision. At the end of the experiment, root primordia were observed only in auxin treated juvenile cuttings. Clustering the expression profiles revealed that the time after excision contributed to expression differences more than the age or auxin. Maximum differences contributed by the age and auxin occurred on day 6 which correlated with the kinetics of root primordia formation. These included genes related to the microtubules (MTs) system. Therefore, expression of 42 transcripts annotated as tubulin or MTs associated proteins (MAPs) was validated in the same RNA samples. The results suggest developmental and auxin regulation of MTs. To determine the relevance to adventitious root (AR) formation, subtle perturbations to MTs were performed with trifluralin during induction. While juvenile cuttings were not affected, improved rooting was obtained in mature cuttings. Taken together it suggests that specific MTs remodeling is required for AR formation in E. grandis. mRNA samlpes from mature and juvenile Eucalyptus sections immediately after cutting served as contorls. The rest of the samples were mature and juvenile sections, treated of not treated with Auxin, 1, 3, 6, 9 and 12 days after the prunning. At each time point, a loop design was performed consisting of a juvenile untreated vs. mature untreated array, a matrue untreated vs. mature auxin treated array, a matrue auxin treated vs. juvenile auxin treated array, and a juvenile auxin treated vs. juvenile untreated array. additional arrays connected between these loops, and between them and the time 0 controls. Most conditions had 3 replicates, but mature-untreated-day1 had 4 replicates, and three conditions had 2 replicates each: Juvenile-auxin-treated-day1, juvenile-untreated-day6, and mature-auxin-treated-day9.

Project description:In order to pinpoint the most differentially expressed genes between Eucalyptus grandis leaf blades and vascular (xylem) tissues as well as between E. grandis and Eucalyptus globulus xylem tissues, a total number of nine 50mer-oligoprobes covering the length of each one of 21,432 unique sequences derived from the Genolyptus EST dataset were synthesized “on-chip” in duplicate, randomly distributed in two blocks of each slide. Probes were also synthesized from ten cDNA sequences encoding known human proteins as negative controls, totaling 21,442 sequences. Leaves and xylem samples were taken from two E. grandis clonal trees, i.e., both derived from the same matrix tree and harboring the same genotype. Two additional xylem samples were collected from two other E. grandis clonal trees of a different genotype, as well as from two E. globulus clonal trees. Therefore, ten cDNA samples and ten identical chips were produced at Roche NimbleGen for the microarray assays, with a total number of 385,956 features per slide. Besides the discovery of differentially expressed genes between leaf and xylem, we wanted to test the validity of the assumed “technical” and “biological duplicates” since all trees were field-grown and four years-old in age. A ten chip study using total RNA recovered from mature leaf and vascular (xylem) tissues of Eucalyptus grandis and xylem from Eucalyptus globulus trees. Two clonal trees of E. grandis (E.grandis_Clone A_Ramet 1 and E.grandis_Clone A_Ramet 2), derived from a single matrix tree and therefore genomically identical, were the source of two samples of leaf RNA and two samples of xylem RNA, individually hybridized to four chips after cDNA synthesis/Cy3 labeling. Two other clonal trees of E. grandis (E.grandis_Clone B_Ramet 1 and E.grandis_Clone B_Ramet 2), derived from a different matrix tree, were the source of two additional samples of xylem RNA individually hybridized to four chips after cDNA synthesis/Cy3 labeling. Likewise, two pairs of clonal trees of E. globulus (E.globulus_Clone A_Ramet 1 and E.globulus_Clone A_Ramet 2/ E.globulus_Clone B_Ramet 1 and E.globulus_Clone B_Ramet 2), derived from two distinct matrix trees, were the source of four additional samples of xylem RNA, individually hybridized to four chips after cDNA synthesis/Cy3 labeling. Each chip measures the expression level of 21,432 genes from Eucalyptus sp. and ten human genes (negative controls) with nine 50-mer probe pairs (PM/MM) per gene in two separate blocks per chip (technical duplicate), totalizing 18 hybridization signal values per gene per chip.

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Expression profiling of PEG induced water stress responsive genes in Eucalyptus grandis

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