ABSTRACT: Two excessive NaCl concentrations (0.25% and 1.5%) and two periods of treatment (7 min and 45 min) were used as moderate- and high-salt sress conditions at early and later stages of response, respectively. As a baseline, samples from cells grown on a standard SIS medium in the same growing system and same illumination were used. Keywords: time course and dose response Cells were grown photosynthetically (10W/m2 illumination) in 60-ml tubes bubbled with 2% CO2, 98% N2. NaCl (dissolved in anoxic buffer) was added at OD600=0.4
Project description:Normal lung function relies on mature function of alveolar type II cels, which have numerous functions including to regulate ion and fluid flux, produce immune molecules, and synthesize and secrete surfactant to stabilize air spaces. Differentiation of type II cells from precursor epithelial cells is accelerated by exposure of cultured cells to glucocorticoid and cAMP. In these studies we used DNA microarray analysis to identify genes of both fetal and adult type II cells that are regulated by glucocorticoid plus cAMP. In the first series of experiments we isolated epithelial cells from fetal human lung and cultured cells for 5 days with or without hormone treatment (dexamethasone plus 8-Br-cAMP plus isobutylmethylxanthine, DCI) and then performed DNA microarray analysis. In the second series of experiments, we isolated type II cells from adult human postmortem lung and cultured cells for 5 days with or without DCI and performed DNA microarray analysis.
Project description:Skeletal muscle has great regenerative capacity, which is dependent on muscle stem cells, also known as satellite cells. We established the purification system of myogenic cells. We used microarrays to identify specific genes during the muscle regenerative process. Myogenic cells were purified at regenerative stages for RNA extraction and hybridization on Affymetrix microarrays. CTX-2d and CTX-5d are the samples derived from injured muscles 2 days and 5 days after cardiotoxin-injection, respectively.
Project description:The expression of genes involved in photosystem development in Rhodobacter sphaeroides is dependent upon three major regulatory networks: FnrL, the PrrBA two-component system and the AppA-PpsR pathway. A PrrA- mutant strain of R. sphaeroides is unable to develop under photosynthetic conditions, but when combined with a ppsR mutation, is able to resume growth. In an effort to better characterize the ppsR regulon and to unravel the relationship between the PrrBA and the AppA-PpsR networks, we compared transcriptome profiles from the wild type, PrrA- and PrrA-PpsR- strains under a permissive, common, growth condition: anaerobic-dark-DMSO. Here is stored the microarray data for the PrrA-PpsR- double mutant strain grown under this condition. Keywords: transcriptome profile Rhodobacter sphaeroides PrrA-PpsR- grown under anaerobic dark DMSO conditions. Sparged with 95% N2, 5% CO2 in sistrom minimal medium A supplemented with final concentration 0.1% YE, 0.5% DMSO. The total RNA from three independent cultures of R. sphaeroides PrrA-PpsR- grown under anaerobic-dark-DMSO conditions was used. cDNA synthesis, fragmentation, labeling and hybridization are described elsewhere (Roh et al., 2004).
Project description:Wild type (2.4.1) strain, dark conditions, controlled oxtgen concentration in the growth medium (achieved by constant sparging with a gas mixture containing 1% CO2, variable - indicated for every sample - concentration O2 and balance N2). Keywords: dose response Cells were harvested at OD 0.16-0.18 after rapid addition of rifampicin, spun down and frozen for subsequent RNA isolation. RNA was isolated using bead beater and RNeasy kit. Traces of DNA were removed by DNAse I treatment until level non-detectable by 33-36 cycles of real-time PCR. Further sample preparaton -according Affymetrix protocols. Data processing: MAS 5.0 values are deposited here (for compatability with other submitted samples), values used in the publications are obtained with RMA and may be recreated using the supplied CEL files.
Project description:We used microarrays to find out carbon souce responsive genes of A. oryzae with 1hr induction with galactose, glucose, and sorbitol. The A.oryzae RIB40 was cultured for 24hr in glucose as a carbon source, then shifted to galactose, glucose, and sorbitol containing medium and cultured for 1hr. Two biological replicates in each conditions were used for the microarray analysis.
Project description:To investigate how histone demethylases KDM4B and KDM6B may be involved in osteogenic commitment of mesenchymal stem cells (MSCs), we performed gene expression profiling and comparison on control, KDM4B- and KDM6B-knockdown MSCs at different stages of osteogenic differentiation. Human MSCs infected with scramble shRNAs, shRNAs against KDM4B or KDM6B are treated with BMP4/7 for 0, 4 and 24hrs. Total RNA were extracted from these 9 samples.
Project description:The goal of this study is to investigate the molecular mechanism of lhx1 on regulation of pronephros formation during the early embryonic development. In the vertebrate embryo the kidney is derived from the intermediate mesoderm. The LIM-class homeobox transcription factor lhx1 is expressed early in the intermediate mesoderm and is one of the first genes to be expressed in the nephric mesenchyme. The animal cap cells can be induced by treatment of activin and retinoic acid to differentiate into pronephros tissue. In this study we investigated the role of Lhx1 in differentiation of pronephros by depleting lhx1 in the organ culture system. We generated the gene expression profile of early pronephros tissue, and demonstrated that expression of genes from all the kidney domains is affected by the absence of lhx1. Taken together our results highlight an essential role for Lhx1 in pronephros formation. lhx1 is involved in driving specification of intermediate mesoderm into nephrogenic mesenchyme. Lhx1 is initially expressed throughout the entire intermediate mesoderm. To determine the role of lhx1 pronephros formation, we performed a microarray analysis using an explant culture system. Xenopus tissue explants can be surgically isolated and cultured under specific conditions to be driven towards many distinct tissue types. Formation of pronephric cell fates is induced by culturing isolated explants in the presence of Activin and RA (AcRA). Treatment of dissected explants of stage 9 blastulae embryos with 10ng/ml Activin and 1x10-4 M retinoic acid can induce differentiation of the pluripotent ectoderm into pan-kidney tissue. For this experiment, both blastomeres of 2-cell embryos were injected with a total of 800pg lhx1 DEED-AS. Explants were dissected and treated with AcRA and expression of pax8 at stage 15 (based on timing of paired control whole embryos) was analyzed. We observed a lack of induction of pax8 expression in lhx1-depleted explants under AcRA treatment conditions in which expression of this gene is normally induced. Based on this observation, microarray analysis was carried out to identify genes whose expression is affected by the absence of lhx1. Explants of injected embryos with 800pg of lhx1 DEED-AS were dissected, treated with pronephric tissue inductive conditions (AcRA) and harvested after 24 hours incubation at 14C (Fig. S5B). The sibling control embryos reached stage 12.5. Explants from uninjected embryos +AcRA and -AcRA as well as explants from DEED injected embryos -AcRA were also harvested. Approximately 12 caps were pooled for each RNA preparation and the analysis was performed using triplicates.
Project description:Our laboratory wanted to define the transcription profile of aged skeletal muscle. For this reason, we performed a triplicate microarray study on young (3 weeks) and aged (24 months) gatrocnemius muscle from wild-type C57B16 Mice Keywords: other this experiment include 2 samples and 6 replicates
Project description:This work presents an exploration of submerged differentiation of the ubiquitous saprophyte and industrially important fungus, Aspergillus niger, in response to a limited availability of a sole carbon and energy source, maltose. In aspergilli and other mold fungi, asexual reproduction through formation of elaborate conidiogenic structures normally requires an aerial interface. This requirement is bypassed in submerged culture in response to severe nutrient limitation. Continuous cultures with cell retention (retentostat cultures) were applied to generate a fundamental physiological state, where the specific growth rate approaches zero, as the density of the cell population adapts to the supply of the limiting energy source. Temporal differentiation of mycelium structure and commitment to asexual reproduction were major phenomena, apparent on biochemical, morphological, physiological, and transcriptomic level. The severe substrate limitation had a rapid negative impact on cytoplasmic processes, and promoted endo- and exogenous nutrient mobilization, and hyphal compartmentalisation. The first conidiogenic structures appeared after one day with little additional differentiation until Day 4 to 6, where a transition to full commitment to reproductive growth took place. Submerged conidiation in A. niger involved transcriptional regulation of homologs of the regulatory pathway, centered around the Bristle gene (brlA), and structural genes previously described in other aspergilli. Comparison of transcriptomes, revealed a number of co-regulated gene clusters, which appear to encode secondary metabolite biosynthetic potential. We discuss the concept of maintenance energy in the context of differentiation, a possible physiological trigger for sporulation and the special physiological adaptations of the starved mycelium. We also present a simple and efficient method for in situ retention of filamentous organisms. The dataset consists of 9 Affymetrix arrays derived from defined growth conditions of lab-scale bioreactor cultures (5L). Total RNA was extracted from biomass harvested at three different growth phases: exponential growth phase, 2 and 8 days of retentostat cultivation. For each of the phases, the data is derived from three biological replicates.
Project description:The root cap surrounds the root meristem, protecting it from harsh environments and facilitating root movement through soil. In Arabidopsis, new root cap cells produced in the meristem displace older cells toward the root periphery, where they undergo programmed cell death and are released from the root. This rapid cell turnover is a unique feature of the root cap and is modulated in part by the combined action of four NAC transcription factors, as well as cell wall modification enzymes. Here we show that the transcription factor NIN-LIKE PROTEIN 7 (NLP7) regulates root cap maturation in Arabidopsis by modulating expression of each of the NAC TFs as well as several cell wall modifying enzymes. NLP7 is a homolog of NIN, a transcription factor required for nodulation in Lotus japonicas, and is highly expressed in the columella root cap. Mutants have altered columella root cap development and decreased levels of homogalacturonan, a major component of pectin. Reverse genetic analysis of genes differentially expressed in nlp7 roots showed that two cell wall modifying enzymes, CELLULASE5 and XTH5, modulate root cap maturation likely downstream of NLP7. Plant cell wall modification is critical for nodulation, and we propose that this characteristic is also present in NLPs. We used microarrays to identify the differences in gene expression between whole roots of WT and nlp7-1 Arabidopsis plants Roots were cut at the root/hypocotyl junction and collected into RLT buffer in the RNeasy plant mini kit. Approximately 60 roots were collected per replicate, with two biological replicates. RNA was extracted using the RNeasy Micro Kit. Probes for array analysis were preparedfrom 1μg total RNA with the one-cycle amplification protocol by Affymetrix according to the manufacturer's instructions. Samples were submitted to Expression Analysis Inc. (Durham, NC) for hybridization to Arabidopsis Whole Genome ATH1 Affymetrix GeneChips.