Transcription profiling of maize pulvinus after gravity treatment
ABSTRACT: Not much is known about the molecular processes involved during gravitropism in monocot plants such as maize. A microarray based study on the expression of genes after a gravity stimulating the maize pulvinus will provide us with valuable information and a better understanding of the underlying molecular processes involved in monocot gravitropism. Objectives for this study included the identification of genes that were regulated at the transcriptional and translation level during gravitropism in the maize pulvinus. This was achieved by microarray analysis of total RNA versus polyribosome associated RNA during a time-course of gravity stimulation of the maize pulvinus. Experiment Overall Design: Six week old maize plants were gravity stimulated by 90º reorientation. Upper (slow elongation) and lower (fast elongation) halves of the most gravity competent pulvini were harvested over a time course ranging from 2 minutes up to one hour (2min, 5 min, 15min, 30min, 60min). Pulvini samples from control (vertical, no gravity stimulation) plants were harvested and labeled as left and right. For each time point, total mRNA and polyribosome-associated mRNA were purified and the transcript profiles analyzed using Affymetrix GeneChip® Maize Genome Arrays. The experiment was repeated twice (two growing seasons) and represent two biological repetitions.
Project description:Not much is known about the molecular processes involved during gravitropism in monocot plants such as maize. A microarray based study on the expression of genes after a gravity stimulating the maize pulvinus will provide us with valuable information and a better understanding of the underlying molecular processes involved in monocot gravitropism. Objectives for this study included the identification of genes that were regulated at the transcriptional and translation level during gravitropism in the maize pulvinus. This was achieved by microarray analysis of total RNA versus polyribosome associated RNA during a time-course of gravity stimulation of the maize pulvinus. Keywords: time course Overall design: Six week old maize plants were gravity stimulated by 90º reorientation. Upper (slow elongation) and lower (fast elongation) halves of the most gravity competent pulvini were harvested over a time course ranging from 2 minutes up to one hour (2min, 5 min, 15min, 30min, 60min). Pulvini samples from control (vertical, no gravity stimulation) plants were harvested and labeled as left and right. For each time point, total mRNA and polyribosome-associated mRNA were purified and the transcript profiles analyzed using Affymetrix GeneChip® Maize Genome Arrays. The experiment was repeated twice (two growing seasons) and represent two biological repetitions.
Project description:osteocyte is the mechanosensor in bone, taking up pivotal position in mediating the mechano-induced bone remodeling. Dimagnetic levitation has been used to stimulating a reduced gravity environment for studing the effects of changed gravity to different organisms.we constructed a superconducting magnet based platform with a large gradient high magnetic field(LG-HMF),which can provide three apparent gravity levels (μ-g,1-g and 2-g). osteocytes are sensetive to gravitational changes, our aim is to explore what responses do osteocytes exists under different gravitational environments in gene level, together with filtering the up- and down-regulated genes. mouse osteocyte-like cell line MLO-Y4 were cultured under three different apparent gravity levels (μ-g,1-g and 2-g) and normal gravity environment (control) for 48 hours, after which total RNA was extracted . And then RNA samples hybridized on affymetrix microarrays to obtain the whole genome expression profiles. the aim that we selected 48 hours as the cell culture time was to make a comparison with our previous researches of osteocytes.
Project description:au13-11_gravity - gravity - Cell cycle and cell proliferation are decoupled under altered gravity conditions. We have previously shown that semisolid cell cultures of Arabidopsis suffer overall genome changes in response to altered gravity and also that cell cycle stages duration is altered. By using synchronized cell cultures we will demonstrate the precise alterations in cell cycle duration and also the transcriptional signature in any of them. - Experiments consists on exposures of Arabidopsis cell cultures to 1g control/simulated microgravity (RPM) conditions. Asynchronous cells exposed for 14 h + Syncronous populations choosen to have an enrichment of cell cycle phases were used (being T7/T10 samples on G2 phase, T14/T16 samples on G1 phase). 6 dye-swap - time course,treated vs untreated comparison
Project description:Genome-wide transcriptional profiling shows that reducing gravity levels in the International Space Station (ISS) causes important alterations in Drosophila gene expression. However, simulation experiments on ground, without space constraints, show weaker effects than space environment. A global and integrative analysis using the “gene expression dynamics inspector” (GEDI) self-organizing maps, reveals a subtle response of the transcriptome using different populations and microgravity and hypergravity simulation devices. These results suggest that, in addition to behavioural responses that can be detected also at the gene expression level, the transcriptome is finely tuned to normal gravity. The alteration of this constant parameter on Earth can have effects on gene expression that depends both on the environmental conditions and the ground based facility used to compensate the gravity vector. Alternative and commons effects of mechanical facilities, like the Random Positioning Machine and a centrifuge, and strong magnetic field ones, like a cryogenically cooled superconductive magnet, are discussed. We compare the effects over the gene expression profile of different gender/age Drosophila imagoes in 3-4 days-long experiments under altered gravity conditions into three GBF ("Ground Based Facilities" for micro/hyper- gravity simulation) using whole genome microarray platforms. Descriptions of different GBFs ("treatments"): LDC means "Large Diameter Centrifuge". Samples can be placed under three conditions: inside LDC (at certain g level), at the LDC rotational control and at external 1g control (outside the LDC). RPM means "Random Positioning Machine". Samples can be placed under two conditions: inside RPM (at nearly 0g, Microgravity level) and at external 1g control (outside the RPM). At the magnet, means INSIDE the Magnetic levitator (another GBF). Samples can be placed under four conditions: inside Magnet 0g* (at microgravity with magnetic field), inside Magnet at 1g* (internal control with magnetic field) or inside the magnet 2g* (at hypergravity with magnetic field) and at external 1g control (outside the magnet)
Project description:Methods: RNA-seq libraries were prepared using the Illumina TruSeq RNA kit and the TrueSeq method was employed for mRNA enrichment. The libraries were quantified and samples were multiplexed in each lane of the flowcell. Cluster generation was performed and then sequenced on the Illumina HiSeq1000 system. Reads were mapped on the Human Genome Reference and normalized expression table was generated. Results: Among differentially expressed genes, 53 of them were up-regulated and 75 were down-regulated. Conclusions: Data demonstrate increased expression of genes associated with growth, development, and pro-survival in cardiac progenitors cultured under simulated microgravity compared with those cultured under standard gravity. RNA-sequencing analysis was performed to compare global gene expression profiles of cells at differentiation day 8 under simulated microgravity vs. standard gravity.
Project description:The basidiomycete Ustilago maydis causes smut disease in maize. Colonization of the host plant is initiated by direct penetration of cuticle and cell wall of maize epidermis cells. The invading hyphae are surrounded by the plant plasma membrane and proliferate within the plant tissue. We identified a novel secreted protein, termed Pep1. Disruption mutants of pep1 are not affected in saprophytic growth and develop normal infection structures. However, Δpep1 mutants fail to penetrate the epidermal cell wall and elicit a strong plant defense response. Using Affymetrix maize arrays we identified about 110 plant genes which are differentially regulated in Δpep1 and wild type infections during the penetration stage. Experiment Overall Design: In three independent experiments plants were infected with the strain SG200Dpep1 which is derived from the solopathogenic U. maydis strain SG200. Samples from infected leaves were taken at 24 hours post infection. Samples were treated under the same conditions as described previosly (Doehlemann et al. (2008) Plant J, in press).
Project description:The classical maize mutant lazy1 (la1), displayed prostrate growth with reduced shoot gravitropism. We compared the transcriptome profile of the third node in la1-ref mutants with those in wild-type plants using RNA-SEQ to examine the genome-wide effect of the ZmLA1 gene. We generated 14.6 and 36.5 million paired-end reads from two biological samples of wild-type and la1-ref mutant plants, respectively.
Project description:Cells from three adult, wild-type, FVB hearts were separated into cardiomyocyte and nonmyocyte fractions using Langendorff perfusion, collagenase digestion and gravity filtration; total RNA was prepared immediately from myocytes, while nonmyocytes were passaged twice to yield a culture of largely fibroblasts from which total RNA was prepared. 6 cardiac polyadenylated RNA (mRNA and lncRNA) and small RNA (microRNA) profiles of isolated cardiomyocytes and fibroblasts from 12-wk FVB/NJ mouse hearts were generated on Illumina HiSeq 2000 instruments.
Project description:Fusarium graminearum gene expression profiles were compared during an early infection time course (1, 2, and 4d post-inoculation) of three monocot hosts (wheat heads, barley heads, and maize developing kernels). Overall design: Fusarium-inoculated vs mock-inoculation, 1d, 2d, and 4d post-inoculation, three biological replicates were done for each time point for each host, 2 technical replicates (dye flips) per sample.
Project description:Diamond Blackfan Anemia (DBA) is associated with developmental defects and profound anemia. Mutations in genes encoding a ribosomal protein of the small (e.g. Rps19) or large (e.g. Rpl11) ribosomal subunit are found in over half of these patients. The mutations cause ribosomal haploinsufficiency, which reduces overall translation efficiency of cellular mRNAs. We reduced expression of *Rps19* or *Rpl11* in mouse erythroblasts and investigated mRNA polyribosome association, which revealed deregulated translation initiation of specific transcripts. Among these were *Bag1*, encoding a Hsp70 co-chaperone, and *Csde1*, encoding an RNA binding protein, both expressed at increased levels in erythroblasts. Their translation initiation is cap-independent and starts from an internal ribosomal entry site (IRES), which appeared sensitive to knock down of Rps19 or Rpl11. Mouse embryos lacking Bag1 die at embryonic day E13.5 with reduced erythroid colony forming cells in the fetal liver, and low Bag1 expression impairs erythroid differentiation in vitro. Reduced expression of Csde1 impairs proliferation and differentiation of erythroid blasts. Protein but not mRNA expression of *BAG1* and *CSDE1* was reduced in erythroblasts cultured from DBA patients. Our data suggest that impaired IRES-mediated translation of mRNAs expressed at increased levels in erythroblasts contributes to the erythroid phenotype of DBA. 3 biological replicates of erythroblasts treated with different shRNA were used for polyribosomal sucrose gradients; RNA was extracted from gradients in 2 samples - mRNA associated with polyribosomes (poly) and the rest (sub).