Expression analysis of the fungus Ustilago maydis WT strain FB2 (a2b2) at different pH values
ABSTRACT: Elicidate the changes of the transcriptional program response of U. maydis WT strain (a2b2) when transfered from neutrality to acidic or alkaline medium The mutants strains CL211 and GP25 used as controls as well as the WT strain are described in Martínez-Espinoza et al., 1997; 2004;González-Prieto et al., 2014 and Banuett & Herskowitz, 1989 respectively. High density-one color arrays from Nimblegen were used according to a design from Scott Gold from University of Georgia. The design has five different oligonucleotides of 60 bp in lenght in duplicate, representing each one of the 6883 gene of the U. maydis genome.
Project description:Alkaline pH triggers an adaptation mechanism in fungi that is mediated by Rim101/PacCp, a zinc finger transcription factor. To identify the genes under its control in Ustilago maydis, we performed microarray analyses, comparing gene expression in a wild type strain vs a rim101/pacC mutant of the fungus. In this study we obtained evidence of the large number of genes regulated mostly directly, but also indirectly (probably through regulation of other transcription factors) by Rim101/PacCp, including proteins involved in a large number of physiological activities of the fungus. Our analyses suggest that the response to alkaline conditions under the control of the Pal/Rim pathway involves changes in the cell wall and plasma membrane through alterations in their lipid, protein, and polysaccharide composition, changes in cell polarity, actin cytoskeleton organization, and budding patterns. Also as expected, adaptation involves regulation by Rim101/PacC of genes involved in the meiotic functions, such as recombination and segregation, and expression of genes involved in ion and nutrient transport, as well as general vacuole functions. The mutant analyzed in this study is described in PMID 15947192 Five different oligonucleotide probes per gene (60 nt in length) per duplicate represented each of the 6,883 genes of the U.maydis genome. Accordingly, the expressed data are equivalent to 10 different assays of the expression of each gene.
Project description:Investigation on expression levels of normal tissue from prostate cancer patients on locus 8q24. 3 chips with 3 arrays each study, using 3 pairs of normal vs. tumor tissue and 3 replicates of the same sample. Each chip contained one pair of normal vs. tumor and one copy of the repeated sample.
Project description:Investigation on expression levels of normal tissue from prostate cancer patients on locus 8q24. The region chr8:127640000-129120000 is tiled with 60 nt probes at 10 nt interval (hg18) 7 chip study, using 7 independent samples.
Project description:Investigation on expression levels of normal tissue from prostate cancer patients on locus 8q24. The region chr8:127640000-129120000 is tiled with isothermal probes (hg17) 7 chip study, using 7 independent samples.
Project description:Embryonic stem cells (ESCs) and induced-pluripotent stem cells (iPSCs) self-renew and differentiate into an array of cell types in vitro and in vivo. A complex network of genetic and epigenetic pathways regulates the self-renewal and differentiation of these pluripotent cells, and the structure and covalent modifications of chromatin play a prominent role in this process. We examine nucleosome occupancy in mouse and human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and differentiated cell types using MNase-seq. To address variability inherent in this technique, we developed a bioinformatic approach that enabled the identification of regions of difference (RoD) in nucleosome occupancy between pluripotent and somatic cells. The majority of changes in nucleosomal signatures that occur in differentiation are reset during reprogramming. We conclude that changes in nucleosome occupancy are a hallmark of pluripotency and likely identify key regulatory regions that play a role in determining cell identity. A six chip study using total RNA recovered from three cell types with 2 replicates each
Project description:Investigation of whole genome gene expression level changes in Sphingomonas. sp A1 AlgO-deficient mutant grown on alginate compared with that on yeast extract AlgO is a possble transcriptional factor described in J. Bacteriol. (2000) 182(14):3998-4004 by Momma K, Okamoto M, Mishima Y, Mori S, Hashimoto W, and Murata K. A two chip study using total RNA recovered from two cultures of Sphingomonas. sp A1 AlgO-deficient mutant grown in 0.5% alginate medium and 0.5% yeast extract medium. Each chip measures the expression level of genes from Sphingomonas. sp A1.
Project description:Investigation of whole genome gene expression level changes of LncRNAs in tumor tissues and paired non-tumor tissues in HBV-positive hapatocellular carcinoma. The different expression genes were further analysised. The human LncRNA microarray analysis of the 10 samples (5 non-tumor tissues and 5 paired tumor tissues) were completed. Total RNA from each sample was quantified using the NanoDrop ND-1000 and RNA integrity was assessed using standard denaturing agarose gel electrophoresis. Total RNA of each sample was used for labeling and array hybridization as the following steps: 1) Reverse transcription with by Invitrogen Superscript ds-cDNA synthesis kit; 2) ds-cDNA labeling with NimbleGen one-color DNA labeling kit; 3) Array hybridization using the NimbleGen Hybridization System and followed by washing with the NimbleGen wash buffer kit; 4) Array scanning using the Agilent Scanner G2505C. Scanned images (TIFF format) were then imported into NimbleScan software (version 2.5) for grid alignment and expression data analysis. Expression data were normalized through quantile normalization and the Robust Multichip Average (RMA) algorithm included in the NimbleScan software. The Probe level (*_norm_RMA.pair) files and mRNA level (*_RMA.calls) files were generated after normalization. All mRNAs level files were imported into Agilent GeneSpring GX software (version 11.5.1) for further analysis.mRNAs that at least 3 out of 6 samples have values greater than or equal to lower cut-off: 50.0 (“All Targets Value”) were chosen for further data analysis. Differentially expressed mRNAs were identified through Volcano Plot filtering. Pathway analysis and GO analysis were applied to determine the roles of these differentially expressed mRNAs played in these biological pathways or GO terms. Finally, Hierarchical Clustering was performed to show the distinguishable mRNAs expression pattern among samples.
Project description:A custom-made microarray was used for comparative transcriptome analysis of transgenic suspension cell line of Nicotiana tabacum that overexpresses the CaRLK1 gene. Ectopic expression of the CaRLK1 preferentially upregulated many amino acid biosynthetic genes under hypoxia: a 1.7-fold higher steady-state concentration of total free amino acids. Free Ala level was predominantly increased by 3-4 times and reached more than half of total free amino acid content. A significantly and markedly increased pyruvate content was also observed. These accumulations are associated with the glyoxylate cycle positively regulated by the CaRLK1. A total of 12 chips were used for microarray. Total RNAs were extracted from BY-2 suspension cells (control), RLK1ox suspension cells (RLK), auxin-free RLK1ox suspension cells (RA), and 10 uM DPI-treated RLK1ox suspension cells (RD) with triple biological replicates.
Project description:A significant percentage of HIV-infected individuals experience a sharp decline in CD4+ T cell counts and progress to AIDS quickly after primary infection. Identification of biomarkers distinguishing rapid progressors (RPs) versus chronic progressors (CPs) is critical for early clinical intervention and could provide novel strategies to facilitate vaccine design and immune therapy. mRNA and miRNA expression profiles in the peripheral blood mononuclear cells (PBMCs) of RPs and CPs were investigated at 111±22 days (Mean±SD) of HIV infection. The association of mRNA and miRNA expression with disease progression was examined by receiver operating characteristic analysis and Kaplan-Meier survival analysis. Pathway enrichment analysis showed that genes with deregulated expression in RPs are primarily involved in apoptosis pathways. Furthermore, we found that 5 miRNAs (miR-31, -200c, -526a, -99a and -503) in RPs were significantly decreased compared to those in CPs (P<0.05). The decreased expression of these miRNAs was associated with rapid disease progression of HIV infection with a 94% predictive value as measured by the area under the curve. The upregulated predicted targets from the 5 signature miRNAs and all upregulated genes identified from mRNA microarray converged to the apoptosis pathway. Moreover, overexpression of miR-31 in primary human T cells promoted their survival. Our results have identified a distinct transcriptomic signature in PBMCs of RPs and provided novel insights to the pathogenesis of HIV infection. A cohort of primary HIV infected individuals with different disease outcome were enrolled in this study. We included 6 individuals with rapid disease progression (RP), seven with chronic disease progression (CP). The HIV infected individuals were never on therapy before the time of sample taken.