Project description:MicroRNAs (miRNAs) play an important role in human brain development and maintenance. To search for miRNAs potentially involved with molecular mechanisms in the pathogenesis of Parkinsons disease (PD), we utilized miRNA microarrays to identify expression changes of 115 annotated Caenorhabditis elegans (C.elegans) miRNAs in human α-synuclein A53T transgenic, dopamine deficient catecholamine transporter gene cat-1 mutant and parkin gene pdr-1 mutant C.elegans strains. Twelve miRNAs were found regulated differentially in α-synuclein transgenic C. elegans, five in cat-1 mutants and three in pdr-1 mutants. The family of miR64/65 appeared co-under-expressed in α-synuclein and cat-1 strains and members of let-7 family co-under-expressed (except miR-241 over-expressed) in a-synuclein and pdr-1 strains. Class H serpentine receptor (srh) family of G-protein-coupled receptor genes were computationally identified to be highly over-represented target candidates for regulated miRNAs.These results indicate that miRNAs are misregulated in C. elegans PD models and suggest a role for these molecules in the disease pathogenesis. Two color NCode microRNA microrarrays were used (Invitrogen) to measure miRNA expression changes in human alpha-synuclein trangenic, cat-1 mutant, and pdr-1 mutant C.elegans strains. The experiment contains four independent biological replicates (worm populations)/strain. Strain N2 was used as reference channel, also prepared from four independently isolated small RNA samples.
Project description:This SuperSeries is composed of the following subset Series: GSE14232: Transcriptome analysis of freshly sorted and expanded regulatory and conventional T cells GSE14233: Detection of differentially methylated regions in CD4+CD25+CD45RA+ regulatory T-cells and conventional CD4+CD25- T-cells GSE14234: Histone H3 Lysine 4 mono-, di- and trimethyl and CTCF in CD4+CD25+CD45RA+ regulatory and conventional CD4+CD25- T-cells Refer to individual Series
Project description:Two reference pools comprised of different miRNA content were hybridized to four different miRNA microarray platforms (Agilent, LC Sciences, Exiqon and a homemade chip (generated from an Inviteogen Ncode probeset)) This dataset includes the reference samples on the Agilent miRNA array. Keywords: miRNA Two reference pools were created from commercially available reference RNA (FirstChoice® mouse Total RNA including the small fraction (Catalogue # AM7800-AM7828, Ambion, Streetsville, ON)). Reference #1 contained equal parts of mouse testicle, ovary and 10-12 day embryo. Reference #2 contained liver, heart and lung. These references were hybridized to four different miRNA microarray platforms (two in two colour, two in one-colour, with 4 replicates each). Data from each platform were normalized by cyclic lowess and analysed by correlation analyses both within and between platforms.
Project description:miRNA expression was compared in skin from control newborn mice and littermate mice ectopically expressing the potent secreted WNT inhibitor Dickkopf 1 (DKK1) in the epidermis. DKK1 completely suppresses hair follicle development. Multiple miRNAs were identified that reproducibly produced signals above background in both types of skin sample. Several miRNAs were identified for which hybridization signals were on average more than 2.5 fold higher in control samples than in Dkk1-expressing samples, suggesting these may be upregulated in hair follicles, and/or are direct or indirect targets of WNT inhibition in the skin. Keywords: miRNA expression array, transgenic mouse, skin, WNT Low molecular weight RNA was isolated using the mirVana RNA extraction kit (Ambion) from full thickness dorsal skin of three K5-rtTA; tetO-Dkk1 newborn mice and three control littermates following doxycycline treatment from E0.5 to induce expression of Dkk1 in double transgenic epidermis [1]. Control and Dkk1âexpressing transgenic samples were tagged for labeling with Cy3 and Cy5 dyes respectively using the Ncode miRNA Labeling System (Invitrogen), mixed by equal mass, and co-hybridized to miRNA arrays that have been described previously [2]. Background-corrected median signals for pixels from each probe spot in both the Cy3 and Cy5 channels were used for analysis. Cy3 (control samples C2, C4, C5) was detected at 532nm and Cy5 (Dkk1âexpressing transgenic samples TG1, TG2, TG4) at 635 nm. References: [1] Chu, E.Y., Hens, J., Andl, T., Kairo, A., Yamaguchi, T.P., Brisken, C., Glick, A., Wysolmerski, J.J., and Millar, S.E. (2004). Canonical WNT signaling promotes mammary placode development and is essential for initiation of mammary gland morphogenesis. Development 131, 4819-4829. [2] Nelson, P.T., Baldwin, D.A., Scearce, L.M., Oberholtzer, J.C., Tobias, J.W., and Mourelatos, Z. (2004). Microarray-based, high-throughput gene expression profiling of microRNAs. Nat Methods 1, 155-161.
Project description:The objective of this study was to identify porcine genes which expression was affected by experimental infection with A. pleuropneumoniae within 24 hours after experimental challenge. Microarray experiments were conducted to reveal genes being significantly differentially expressed in infected versus non-infected lung tissue and in liver and lung lymph node tissue from three challenged versus two non-challenged animals. Keywords: Host response Three comparisons were done: - infected versus non-infected lung tissue sampled from three challenged pigs - liver tissue from three challenged versus two non-challenged animals - lung lymph node tissue from three challenged versus two non-challenged animals All experiments were conducted as common reference design.
Project description:Analysis of Histone H3 Lysine 4 mono-, di- and trimethyl and the boundary protein CTCF in CD4+CD25+CD45RA+ regulatory T-cells and conventional CD4+CD25- T-cells. To investigate regulatory functions or potential new transcription start sites in Treg and Tconv cells, we investigated the associated histone modifications. Mono- and dimethylation of histone 3 lysin 4 (H3K4) were previously shown to mark enhancer regions, whereas H3K4 trimethylation generally associates with transcription start sites. At imprinted loci, binding of the insulator protein CTCF, which restricts or directs enhancer-promoter interactions, is often regulated by DNA-methylation. Therefore we performed ChIP-on-chip experiments (chromatin immunoprecipitation followed by microarray hybridization; samples were amplified with ligation mediated PCR [see label protocol for the procedure] prior to labeling) for mono- di- and trimethylation of histone 3 lysin 4 and of CTCF in expanded Treg and Tconv cells. Keywords: ChIP-on-chip ChIP-on-chip experiments for H3K4 mono-, di- and trimethyl and CTCF in CD4+CD25+CD45RA+ regulatory T-cells and conventional CD4+CD25- T-cells were co-hybridizied with the input. Three biologiacal replicates (rep1-3) were performed for every histone mark, two CTCF (rep1 and rep2).
Project description:We have previously developed an approach that fractionates genomic DNA fragments depending on their CpG density (methyl-CpG-immunoprecipitation, MCIp), and adapted this approach to identify regions that are differentially methylated in the two closely related regulatory T-cells (Treg cells) and conventional T-cells (Tconv cells). Because Treg cells naturally occur at a relatively low frequency, we used a previously established protocol to expand Treg cells from a stable naïve Treg population that is characterized by the co-expression of CD4, CD25 and CD45RA. We separated gDNA of both expanded T cell lineages (Tregexp and Tconvexp) into unmethylated (CpG) and methylated pools (mCpG) using MCIp and compared cell type-specific differences in DNA methylation by co-hybridization of the two umethylated or the two methylated DNA subpopulations of Treg and Tconv, respectively, to these locus-wide custom tiling arrays. As enriched DNA-fragments from a cell type in the methylated fraction should be depleted in the unmethylated fraction, the signal intensities in CpG pool and mCpG pool hybridizations should complement themselves (âMirror-Imageâ approach) and thereby allow the identification of differentially methylated regions (DMR). Because we expected to find lineage-specific methylation differences with greater probability in regions associated with differential transcriptional activity, we limited our analysis to gene loci that showed cell type-specific gene expression in Treg versus Tconv cells plus a handful of control regions that were equally expressed in both cell types. The microarray used in this study covered 12 megabases of the human genome and contained 69 regions (with a median size of 100.000 kb) and 128 proximal promoter regions and 181 genes, which included a number of well known and functionally relevant genes like CD40LG, IFNG, FOXP3, IL2RA and CTLA4. Keywords: MCIp-on-chip; comparative genomic hybridization With MCIp gDNA from Treg or Tconv cells was separated into hypo- and hypermethylated pools. On each array, wheather the two hypomethylated fractions- one from Treg, the other from Tconv cells- or the two hypermethylated fractions were cohybridized. Two biological replicates.
Project description:To globally define methylation-âproneâ and -âprotectedâ CpG islands in leukemia, we analyzed the methylation status of 23,000 CpG islands of the human genome in two acute leukemia cell lines as well as normal blood monocytes using our previously described methyl-CpG immunoprecipitation (MCIp) technique (Gebhard et al. 2006; Schilling and Rehli 2007). This method enriches for highly CpG methylated DNA that can be directly applied to fluorescent labeling and oligonucleotide microarray hybridization without an additional amplification step. Keywords: MCIp-on-Chip; comparative genomic hybridization CpG-methylated genomic DNA was enriched using methyl-CpG immunoprecipitation (MCIp). On each microarray, the enriched material from leukemia cell lines was compared to the enriched material from normal blood monocytes to identify aberrantly methylated regions. Three biological replicates were analysed for each cell line.
Project description:To study the correlation between sequence motif appearance, transcription factor binding and aberrant hypermethylation in the cell lines, we performed ChIP-on-chip analyses (on CpG island microarrays) for the transcription factors Sp1, NRF1 and YY1 in normal peripheral blood monocytes. Keywords: ChIP-on-Chip; comparative genomic hybridization Transcription factor bound genomic DNA was enriched using ChIP. On each microarray, the enriched material was compared to the genomic input to identify transcription factor bound regions. Two biological replicates were analysed for each factor.
Project description:Global transcriptional responses in duodenal intestinal epithelia of chickens following primary and secondary Eimeria acervulina infections. Simple loop hybridization (day 0 vs. days 1-2, days 1-2 vs. 3-4, days 3-4 vs. 5-6, and days 5-6 vs. 7-8) per primary or secondary infections.