Journal: Biomaterials

Article Title: Genomic analysis of the role of transcription factor C/EBP? in the regulation of cell behaviour on nanometric grooves

doi: 10.1016/j.biomaterials.2012.11.036

Figure Lengend Snippet: Results of C/EBPβ and C/EBPδ ChIP-SEQ and microarray. a) MEF WT cells were incubated in the presence or absence of a combination of the cyclic AMP-elevating agents, forskolin and rolipram (F/R). Following stimulation cells were fixed, chromatin was isolated and then immunoprecipitated (ChIP'd) with anti-C/EBPβ or C/EBPδ antibodies, as described in Materials and methods. ChIP'd DNA samples were then sequenced (ChIP-SEQ) on an Illumina GA IIx DNA sequencer. ChIP analysis of the resulting DNA sequences was then performed using the Homer (version 3.9) suite of tools ( biowhat.ucsd.edu/homer/ngs/index.html ). The figure shows part of the Homer analysis indicating that aligned sequences from each ChIP experiments contained bone fide C/EBP consensus binding motifs, hence validating the experimental technique. b) MEF WT, MEF C/EBPβ −/− and MEF C/EBPδ −/− cells were stimulated for 5 h in the presence or absence of F/R. Cells were then harvested, RNA isolated, biotinylated sense-strand DNA synthesised and hybridised to mouse whole genome GeneChip ® ST Arrays (Affymatrix). A Venn diagram was then generated to illustrate the degree of overlap between genes identified by ChIP-SEQ as containing consensus, F/R-dependent C/EBP-binding sites and F/R-induced changes in MEF gene expression.

Article Snippet: Briefly, reverse transcription was used to prime poly(A) and non-poly(A), but not ribosomal, mRNA and generate sense strand cDNA for fragmentation and labelling, using the Affymetrix GeneChip® WT Terminal Labelling Kit (PN 900671).

Techniques: Chromatin Immunoprecipitation, Microarray, Incubation, Isolation, Immunoprecipitation, Next-Generation Sequencing, Binding Assay, Generated, Expressing

Journal: Molecular Neurodegeneration

Article Title: Decreased expression of B cell related genes in leukocytes of women with Parkinson's disease

doi: 10.1186/1750-1326-6-66

Figure Lengend Snippet: Significant expression changes in peripheral blood leukocytes of Parkinson's disease patients compared to controls . ( A ) Schematic representation of the statistical analysis and the filters applied on the Affymetrix GeneChip Human Exon 1.0 ST Array exon-level expression data. The numbers of exon-level probesets remained after each filtering step are indicated in circles. ( B ) PCA mapping of patients and control samples according to the expression levels of the 206 significantly changed exon probesets.

Article Snippet: GeneChip whole-transcript sense target labeling of 1 μg of total RNA was done according to the manufacturer's instructions (GeneChip WT cDNA Synthesis and Amplification and GeneChip WT Terminal Labeling Kits, Affymetrix Inc., Santa Clara, CA).

Techniques: Expressing

Journal: Oncotarget

Article Title: Characterization of a P-Rex1 gene signature in breast cancer cells

doi: 10.18632/oncotarget.10285

Figure Lengend Snippet: Changes in gene expression induced by HRG in T-47D cells T-47D cells were treated with HRG (20 ng/ml) or vehicle for 6 h. Total RNA from three replicates was extracted and reverse transcribed to cDNA. Gene expression profiling was carried out using an Affymetrix GeneChip Human Gene 1.0 ST Array. ( A ) Heatmap of the microarray data showing changes in gene expression by HRG ( p -values

Article Snippet: Total RNA for each sample was obtained using the miRNeasy kit (Qiagen; Valencia, CA), reversed transcribed to cDNA (GeneChip® Whole Transcript cDNA Synthesis and Amplification Kit, Affymetrix; Santa Clara, CA), fragmented, and end labeled (GeneChip® WT Terminal Labelling Kit, Affymetrix) for further hybridization to an Affymetrix GeneChip Human Gene 1.0 ST Array (GPL6244) according to the standard protocols at the University of Pennsylvania Molecular Profiling Facility.

Techniques: Expressing, Microarray

Journal: Oncotarget

Article Title: Characterization of a P-Rex1 gene signature in breast cancer cells

doi: 10.18632/oncotarget.10285

Figure Lengend Snippet: Effect of P-Rex1 RNAi on the expression of genes regulated by HRG T-47D cells were transfected with two different P-Rex1 RNAi sequences ( P-Rex #1 and P-Rex #2 ), or a non-target control RNAi ( NTC ). After 16 h, cells were serum starved for 48 h and stimulated with HRG (20 ng/ml) or vehicle for 6 h. Gene expression profiling was carried out using an Affymetrix GeneChip Human Gene 1.0 ST Array. P-Rex1-regulated genes were defined as those in which both P-Rex1 RNAi duplexes (#1 and #2) caused a statistically significant change ( p

Article Snippet: Total RNA for each sample was obtained using the miRNeasy kit (Qiagen; Valencia, CA), reversed transcribed to cDNA (GeneChip® Whole Transcript cDNA Synthesis and Amplification Kit, Affymetrix; Santa Clara, CA), fragmented, and end labeled (GeneChip® WT Terminal Labelling Kit, Affymetrix) for further hybridization to an Affymetrix GeneChip Human Gene 1.0 ST Array (GPL6244) according to the standard protocols at the University of Pennsylvania Molecular Profiling Facility.

Techniques: Expressing, Transfection

Journal: Veterinary Research

Article Title: Chicken interferome: avian interferon-stimulated genes identified by microarray and RNA-seq of primary chick embryo fibroblasts treated with a chicken type I interferon (IFN-α)

doi: 10.1186/s13567-016-0363-8

Figure Lengend Snippet: Correlation of ISGs identified as significant by RNA-seq and microarray. Venn diagrams showing correlation of significant ISGs (FC ≥ 3; FDR ≤ 0.01, unless stated otherwise) for: (i) Illumina 100b paired-end RNA-seq, (ii) Affymetrix 32K GeneChip Chicken Genome Array and (iii) Chicken Gene 1.0 ST Array, following induction of CEF for 6 h with 1000 units of rChIFN1. ( A ) Total hits (“ n =”) shown for each technology; those corresponding to Galgal4 assembly Gene IDs are shown in brackets (“recognised”)—RNA-seq hits all represent Galgal4 mapped genes. ( B ) Hits from array technologies were manually curated to maximise numbers of corresponding genes. ( C ) Curated array hits shown in ( B ) that are present amongst RNA-seq hits, but at lower levels of significance, were transferred to the respective RNA-seq-overlapping sectors. Total genes are shown for ( A – C ).

Article Snippet: RNA samples were processed for microarray with the GeneChip® Chicken Genome Array (Affymetrix) using the GeneChip® 3′ IVT Express Kit (Affymetrix) or for microarray with the Chicken Gene 1.0 ST Array (Affymetrix) using the Ambion (Paisley, UK) WT Expression Kit for Affymetrix GeneChip® Whole Transcript (WT) Expression Arrays (Ambion) and the GeneChip WT Terminal Labelling and Controls Kit (Ambion), following the manufacturers’ instructions, as described previously [ ].

Techniques: RNA Sequencing Assay, Microarray

Journal: Veterinary Research

Article Title: Chicken interferome: avian interferon-stimulated genes identified by microarray and RNA-seq of primary chick embryo fibroblasts treated with a chicken type I interferon (IFN-α)

doi: 10.1186/s13567-016-0363-8

Figure Lengend Snippet: Comparison of expression level and rank of significant ISGs identified by RNA-seq and microarrays. Spearman correlation plots for significant ISGs from: (i) Illumina 100b paired-end RNA-seq and (ii) Affymetrix 32K GeneChip Chicken Genome Array, following induction of CEF for 6 h with 1000 units of rChIFN1, by FC ( A ) and by Rank ( B ).

Article Snippet: RNA samples were processed for microarray with the GeneChip® Chicken Genome Array (Affymetrix) using the GeneChip® 3′ IVT Express Kit (Affymetrix) or for microarray with the Chicken Gene 1.0 ST Array (Affymetrix) using the Ambion (Paisley, UK) WT Expression Kit for Affymetrix GeneChip® Whole Transcript (WT) Expression Arrays (Ambion) and the GeneChip WT Terminal Labelling and Controls Kit (Ambion), following the manufacturers’ instructions, as described previously [ ].

Techniques: Expressing, RNA Sequencing Assay

Journal: Veterinary Research

Article Title: Chicken interferome: avian interferon-stimulated genes identified by microarray and RNA-seq of primary chick embryo fibroblasts treated with a chicken type I interferon (IFN-α)

doi: 10.1186/s13567-016-0363-8

Figure Lengend Snippet: Correlation of ISGs identified as significant by RNA-seq and microarray. Venn diagrams showing correlation of significant ISGs (FC ≥ 3; FDR ≤ 0.01, unless stated otherwise) for: (i) Illumina 100b paired-end RNA-seq, (ii) Affymetrix 32K GeneChip Chicken Genome Array and (iii) Chicken Gene 1.0 ST Array, following induction of CEF for 6 h with 1000 units of rChIFN1. ( A ) Total hits (“ n =”) shown for each technology; those corresponding to Galgal4 assembly Gene IDs are shown in brackets (“recognised”)—RNA-seq hits all represent Galgal4 mapped genes. ( B ) Hits from array technologies were manually curated to maximise numbers of corresponding genes. ( C ) Curated array hits shown in ( B ) that are present amongst RNA-seq hits, but at lower levels of significance, were transferred to the respective RNA-seq-overlapping sectors. Total genes are shown for ( A – C ).

Article Snippet: RNA samples were processed for microarray with the GeneChip® Chicken Genome Array (Affymetrix) using the GeneChip® 3′ IVT Express Kit (Affymetrix) or for microarray with the Chicken Gene 1.0 ST Array (Affymetrix) using the Ambion (Paisley, UK) WT Expression Kit for Affymetrix GeneChip® Whole Transcript (WT) Expression Arrays (Ambion) and the GeneChip WT Terminal Labelling and Controls Kit (Ambion), following the manufacturers’ instructions, as described previously [ ].

Techniques: RNA Sequencing Assay, Microarray

Journal: Veterinary Research

Article Title: Chicken interferome: avian interferon-stimulated genes identified by microarray and RNA-seq of primary chick embryo fibroblasts treated with a chicken type I interferon (IFN-α)

doi: 10.1186/s13567-016-0363-8

Figure Lengend Snippet: Comparison of expression level and rank of significant ISGs identified by RNA-seq and microarrays. Spearman correlation plots for significant ISGs from: (i) Illumina 100b paired-end RNA-seq and (ii) Affymetrix 32K GeneChip Chicken Genome Array, following induction of CEF for 6 h with 1000 units of rChIFN1, by FC ( A ) and by Rank ( B ).

Article Snippet: RNA samples were processed for microarray with the GeneChip® Chicken Genome Array (Affymetrix) using the GeneChip® 3′ IVT Express Kit (Affymetrix) or for microarray with the Chicken Gene 1.0 ST Array (Affymetrix) using the Ambion (Paisley, UK) WT Expression Kit for Affymetrix GeneChip® Whole Transcript (WT) Expression Arrays (Ambion) and the GeneChip WT Terminal Labelling and Controls Kit (Ambion), following the manufacturers’ instructions, as described previously [ ].

Techniques: Expressing, RNA Sequencing Assay

Journal: PLoS ONE

Article Title: Multiple Roles of Integrin-Linked Kinase in Epidermal Development, Maturation and Pigmentation Revealed by Molecular Profiling

doi: 10.1371/journal.pone.0036704

Figure Lengend Snippet: Targeted inactivation of Ilk and loss of ILK transcripts and protein in mouse epidermis. ( A ) The skin of 3 day-old K14Cre-Ilk f/+ and K14Cre-Ilk f/f mice (4 each, from two different litters) was isolated and treated with dispase to separate the epidermis from the dermis. Epidermal lysates were prepared and analyzed by immunoblot with antibodies against ILK or γ-tubulin, used as loading control. ( B ) Total RNA from K14Cre-Ilk f/+ and K14Cre-Ilk f/f mice (5 each) was used to interrogate GeneChip Mouse Gene 1.0 ST Arrays, and the signal intensity corresponding to individual Ilk exons was analyzed. The average intensity values obtained for each exon in RNA from K14Cre-Ilk f/f epidermis are shown relative to those in K14Cre-Ilk f/+ tissues, which have been set to 1.0.

Article Snippet: For probe preparation, cDNA synthesis and in vitro transcription from 5 µg total RNA were done with an Ambion WT Expression Kit for Affimetrix GeneChip Whole Transcript WT Expression Arrays (Ambion, Foster City, CA), as per manufacturer's protocols.

Techniques: Mouse Assay, Isolation

Journal: PLoS Genetics

Article Title: p63 exerts spatio-temporal control of palatal epithelial cell fate to prevent cleft palate

doi: 10.1371/journal.pgen.1006828

Figure Lengend Snippet: p63 regulates an adhesion programme in the secondary palate. ( A ) Heatmap of genes encoding proteins involved in cell adhesion that are differentially-expressed in the palatal shelves of wild-type versus p63 -/- embryos (green, down-regulation; black, no significant change; red, up-regulation). ( B ) Functional annotation of differentially-expressed genes in the palatal shelves of wild-type versus p63 -/- embryos as assessed by DAVID analysis. ( C ) qPCR analysis of genes indicated as differentially-regulated in microarray analyses using RNA extracted from palatal shelves dissected from E14.0 wild-type and p63 -/- mice. ( D ) ChIP-qPCR analysis of p63-bound sites within 25 kb of cell adhesion genes. Fold-enrichment for each binding region was calculated relative to a control region in exon 2 of myoglobin (set at 1; pale bar), to which p63 does not bind. Asterisks represent level of significance: * = P

Article Snippet: Microarray analysis Amplified sense-strand cDNA was generated from 100 ng of total RNA (Ambion WT Expression Kit).

Techniques: Functional Assay, Real-time Polymerase Chain Reaction, Microarray, Mouse Assay, Chromatin Immunoprecipitation, Binding Assay