Journal: Nucleic Acids Research

Article Title: The Effects of Sequence Variation on Genome-wide NRF2 Binding—New Target Genes and Regulatory SNPs

doi: 10.1093/nar/gkw052

Figure Lengend Snippet: Testing the limits of NRF2 binding. Systematic multivariate ARE results by protein binding microarray for NRF2-MAFG heterodimer are shown. Tolerance for sequence variation was studied for positions 1–3 (white), 6–8 (grey) and 9–11 (dark grey) ( A ) and positions 1, 5 and 11 (seed sequences are shown in grey) ( B ). Results are depicted as measured binding relative to NQO1.ARE binding (black) (mean ± S.E.M., n = 45).

Article Snippet: The dilutions were dispensed in a 384-well plate (polypropylene plate No 267462, Nunc, N.Y, USA) and printed onto the avidin-coated glass slides with a microarray printer (BioRobotics MicroGrid II, BioRobotics Ltd, Cambridge, UK).

Techniques: Binding Assay, Protein Binding, Microarray, Sequencing

Journal: Nucleic Acids Research

Article Title: The Effects of Sequence Variation on Genome-wide NRF2 Binding—New Target Genes and Regulatory SNPs

doi: 10.1093/nar/gkw052

Figure Lengend Snippet: A SNP in FTL promoter has drastic effects of NRF2 binding and transcriptional activation. ( A ) A promoter analysis of the FTL gene at chr19 showing the location of experimentally verified NRF2 binding ARE together with dbSNP (v138) and ENCODE ChIP-seq data. ChIP-seq track displays combined MAFF and MAFK binding signals in H1-hESC (MAFK), K562 (MAFF, MAFK), HeLa-S3 (MAFK), HepG (MAFF, MAFK) and IMR90 (MAFK) cell lines. ( B ) Detailed view showing FTL ARE sequence and the SNP (rs113067944, A→C) position. ( C ) Protein binding microarray results for FTL.ARE.A and the SNP bearing FTL.ARE.C. Results are calculated as measured binding relative to NQO1.ARE binding (mean ± S.E.M, n = 39.). Scramble oligonucleotides served as negative control. ( D ) HEK-293T cells were transfected with NQO1-ARE and FTL-ARE bearing either allele A or allele C with and without NRF2 -expressing plasmids. Twenty-four h after transfection cells were treated with NRF2 inducer (L-SFN) for 16 h followed by luciferase activity measurements. An empty pGL3 promoter vector served as control and activities were normalized to β-galactosidase activity. Results are shown relative to control (mean± S.E.M, n = 4).

Article Snippet: The dilutions were dispensed in a 384-well plate (polypropylene plate No 267462, Nunc, N.Y, USA) and printed onto the avidin-coated glass slides with a microarray printer (BioRobotics MicroGrid II, BioRobotics Ltd, Cambridge, UK).

Techniques: Binding Assay, Activation Assay, ChIP-sequencing, Sequencing, Protein Binding, Microarray, Negative Control, Transfection, Expressing, Luciferase, Activity Assay, Plasmid Preparation, Control

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: Self-self hybridization at 56°C of 5 μ g (per dye) of labeled total mouse brain RNA. Diagrams of average intensity values show filtered results in logarithmic scale. The y axis represents values of AlexaFluor3 dye measurement at 532 nm, and the x axis represents values of AlexaFluor5 dye measurement at 635 nm. Red spots represent all signals from LNA and DNA probes spotted on the microarray slide. (a, c) Detection of MiR-9 (yellow spots, black arrow) and miR-9* (yellow spots, white arrow) with LNA probes. (b, d) Detection of snoRNAs (yellow spots) with antisense DNA probes (including mismatch probes in (d)). Sense DNA probes signals were below detection levels and thus filtered out. (a, b) RNA labeling using the protocol described by manufacturer. (c, d) Modified RNA labeling (see ).

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Hybridization, Labeling, Microarray, Modification

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: Self-self hybridization at 56°C of 1 μ g (per dye) of labeled total mouse brain RNA. Diagrams of average intensity values show filtered results in logarithmic scale. The y axis represents values of AlexaFluor 3 dye measurement at 532 nm, and the x axis represents values of AlexaFluor5 dye measurement at 635 nm. Red spots represent all signals from LNA and DNA probes spotted on the microarray slide. Detection of (a) tRNAs (yellow spots) and (b) 7SK RNA (yellow spots) with antisense DNA probes (including mismatch and deletion probes). Sense DNA probes were below detection levels and thus filtered out. (c) Diagram showing mean intensity values ( y axis) of DNA probes for detection of highly structured ncRNAs.

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Hybridization, Labeling, Microarray

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: Self-self hybridization at 64°C of 2 μ g (per dye) of labeled total mouse brain RNA. Diagrams of average intensity values show filtered results in logarithmic scale. The y axis represents values of AlexaFluor3 dye measurement at 532 nm, and the x axis represents values of AlexaFluor5 dye measurement at 635 nm. Red spots represent all signals from LNA and DNA probes spotted on the microarray slide. (a) Detection of snoRNAs (yellow spots) with antisense DNA probes. (b) Detection of miR-9 (yellow spots, black arrows) and miR-9* (yellow spots, white arrow). Sense DNA probes were below detection levels and filtered out. (c) Diagram showing mean intensity values ( y axis) of all antisense DNA probes detecting snoRNAs.

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Hybridization, Labeling, Microarray

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: Self-self hybridization at 64°C of 2 μ g (per dye) of labeled total mouse brain RNA. Diagrams of average intensity values show filtered results in logarithmic scale. The y axis represents values of AlexaFluor3 dye measurement at 532 nm, and the x axis represents values of AlexaFluor5 dye measurement at 635 nm. Red spots represent all signals from LNA and DNA probes spotted on the microarray slide. (a) Detection of snoZ39 (yellow spots) with antisense DNA probes (black arrow, blow up) and one nucleotide mismatch DNA probe (white arrow, blow up). (b) Diagram showing on the y axis the percentage of mean detection value of the antisense probe SNOZ39−6–60MM1 and the one nucleotide mismatch probe SNOZ39−6–60MM1. (c) Diagram showing the mean intensity values ( y axis) of the antisense probes of one (MM1) and two (MM2) nucleotide mismatch probes for 7SK RNA and SNORD55.

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Hybridization, Labeling, Microarray

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: (a) Heat map showing differential expression between brain and mouse embryonic stem cells of ncRNAs spotted on the DNA-LNA microarray. Up- and downregulation of ncRNAs in brain are indicated with red or green color, respectively. Only differential expression of at least two folds is indicated. (b) Northern blot showing expression of SNORD55 and SNORA71 in mouse embryonic stem cells and mouse brain. Ten micrograms of total RNA were used, and 5.8S rRNA was used as a loading control.

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Quantitative Proteomics, Microarray, Northern Blot, Expressing, Control

Journal: Journal of Nucleic Acids

Article Title: Expression Profiling of a Heterogeneous Population of ncRNAs Employing a Mixed DNA/LNA Microarray

doi: 10.1155/2012/283560

Figure Lengend Snippet: Real-time PCR verification of differential expression of selected ncRNAs captured by DNA and LNA probes on the DNA/LNA microarray platform. Results are represented as relative expression levels between mouse ES cells and mouse brain. Data are shown as mean ± SEM; n = 5; * P < 0.05; ** P < 0.01, *** P < 0.005 significantly different from mES cells by Student t -test.

Article Snippet: The LNA-based capture probe set for short ncRNAs as well as the self-designed DNA-based capture probe set for long ncRNAs was spotted on HiSens epoxy-coated glass slides (Nexterion) using the MicroGrid II Microarray Spotter (Zinsser Analytic).

Techniques: Real-time Polymerase Chain Reaction, Quantitative Proteomics, Microarray, Expressing