Dataset Information

EV23+24 mRNA levels in Wild-type versus ddm1/+ backcross bolting Arabidopsis thaliana plants

ABSTRACT: The two samples in this series are complementary hybridizations in a dye-swap analysis. Below is the normalized result of the paired dye swap samples EV23 and EV24. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using both family-wise error rate (FWER) and false discovery rate (FDR) controlling methods. The step-down multiple comparisons procedure of Holm (1979) was used to control the FWER below alpha = 0.01, while the step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found to be significantly enriched for DNA methylation after hypothesis-testing with a controlled error rate are flagged in the last two columns. Keywords: other

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE1324 | GEO | 2004/06/12

SECONDARY ACCESSION(S): PRJNA91025

REPOSITORIES: GEO

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Project description:The purpose of the McrBC methylation microarray assay is to determine which regions of a genome are methylated versus those that are unmethylated in a single Arabidopsis thaliana genotype. McrBC is a methylation-sensitive enzyme that restricts DNA only at purine-Cmethyl half sites when separated between 50bp and 3kb. A designated amount of DNA from a particular genotype is sheared to a size range of 1kb-10kb using nebulization. We restrict half of the nebulized DNA with McrBC, and the methylated fraction is then removed from the unmethylated fraction through gel purification of DNA fragments greater than 1kb.* The remaining nebulized DNA is subjected to the same gel purification scheme, but with no McrBC treatment. In a single hybridization, the untreated sample is labeled with Cy5 and the McrBC-treated sample with Cy3. Thus, after labeling and microarray hybridization, the ratio of normalized Cy5 to normalized Cy3 represents the relative methylation at the sequence represented by the spot on the microarray. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. ddm1 seedlings, 9 days old. The two samples in this series are complementary hybridizations in a dye-swap analysis. This is the normalized result of the paired dye swap samples VC131 and VC135. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using both family-wise error rate (FWER) and false discovery rate (FDR) controlling methods. The step-down multiple comparisons procedure of Holm (1979) was used to control the FWER below alpha = 0.01, while the step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found to be significantly enriched for DNA methylation after hypothesis-testing with a controlled error rate are flagged in the last two columns. Keywords: other

Project description:The purpose of the McrBC methylation microarray* assay is to determine which regions of a genome are methylated versus those that are unmethylated in a single Arabidopsis thanliana genotype. McrBC is a methylation-sensitive enzyme that restricts DNA only at purine-Cmethyl half sites when separated between* 50bp and 3kb. A designated amount of DNA from a particular genotype is sheared to a size range of 1kb-10kb using nebulization. We restrict half of the nebulized DNA with McrBC, and the methylated fraction is then removed from the unmethylated fraction through gel purification of DNA fragments greater than 1kb.* The remaining nebulized DNA is subjected to the same gel purification scheme, but with no McrBC treatment. In a single hybridization, the untreated sample is labeled with Cy5 and the McrBC-treated sample with Cy3.* Thus, after labeling and microarray hybridization, the ratio of normalized Cy5 to normalized Cy3 represents the relative methylation at the sequence represented by the spot on the microarray.* Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis. wild-type seedlings, 9 days old. Below is the normalized result of the paired dye swap samples VC134 and VC136. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using both family-wise error rate (FWER) and false discovery rate (FDR) controlling methods. The step-down multiple comparisons procedure of Holm (1979) was used to control the FWER below alpha = 0.01, while the step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found to be significantly enriched for DNA methylation after hypothesis-testing with a controlled error rate are flagged in the last two columns. Keywords: other

Project description:This series is a technical replicate of VC133+135. The purpose of the McrBC methylation microarray assay is to determine which regions of a genome are methylated versus those that are unmethylated in a single Arabidopsis thanliana genotype. McrBC is a methylation-sensitive enzyme that restricts DNA only at purine-Cmethyl half sites when separated between 50bp and 3kb. A designated amount of DNA from a particular genotype is sheared to a size range of 1kb-10kb using nebulization. We restrict half of the nebulized DNA with McrBC, and the methylated fraction is then removed from the unmethylated fraction through gel purification of DNA fragments greater than 1kb. The remaining nebulized DNA is subjected to the same gel purification scheme, but with no McrBC treatment. In a single hybridization, the untreated sample is labeled with Cy5 and the McrBC-treated sample with Cy3. Thus, after labeling and microarray hybridization, the ratio of normalized Cy5 to normalized Cy3 represents the relative methylation at the sequence represented by the spot on the microarray. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis. ddm1 seedlings, 9 days old. Below is the normalized result of the paired dye swap samples VC133 and VC137. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using both family-wise error rate (FWER) and false discovery rate (FDR) controlling methods. The step-down multiple comparisons procedure of Holm (1979) was used to control the FWER below alpha = 0.01, while the step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found to be significantly enriched for DNA methylation after hypothesis-testing with a controlled error rate are flagged in the last two columns. Keywords: other

Project description:This series is a biological replicate of EV55+56 The purpose of the chromatin immunoprecipitation/microarray (ChIP/chip) experiment is to determine which regions of a genome are enriched for a particular histone modification in a single Arabidopsis thanliana genotype. Chromatin immunoprecipitation with antibodies raised against dimethyl histone-H3 lysine-9 (H3mK9) or dimethyl histone-H3 lysine-4 (H3mK4) is performed on a selected genotype. This purified DNA from each immunoprecipiation (mH3K9, mH3K4, no antibody control) is used for random amplification to increase the quantity of DNA for microarray hybridization. The amplified DNA from each experimental sample is then labeled with Cy5 and hybridized against total input DNA from the corresponding genotype, labeled in Cy3. In a single hybridization, the total input DNA serves as a baseline and is compared to the immunoprecipitated samples. Ratios of normalized signal intensities were calculated to identify enrichment of a particular sequence after immunoprecipitation, in comparison to the total input DNA. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis These data were normalized and subjected to hypothesis testing. Error rate was controlled only by Benjamini and Hochberg's step-up procedure for limiting the False Discovery Rate. Ddm1 seedlings, 9 days old. This is the normalized result of the paired dye swap samples EV104 and EV105. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using false discovery rate (FDR) controlling methods. The step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found after hypothesis-testing with a controlled error rate to be significantly enriched or depleted for H3K4 methylation compared to mean values found in euchromatic regions are flagged in the FDR column. No family-wise error rate method was used to analyze this sample. Keywords: other

Project description:This series is a biological replicate of EV49+50. The purpose of the chromatin immunoprecipitation/microarray (ChIP/chip) experiment is to determine which regions of a genome are enriched for a particular histone modification in a single Arabidopsis thanliana genotype. Chromatin immunoprecipitation with antibodies raised against dimethyl histone-H3 lysine-9 (H3mK9) or dimethyl histone-H3 lysine-4 (H3mK4) is performed on a selected genotype. This purified DNA from each immunoprecipiation (mH3K9, mH3K4, no antibody control) is used for random amplification to increase the quantity of DNA for microarray hybridization. The amplified DNA from each experimental sample is then labeled with Cy5 and hybridized against total input DNA from the corresponding genotype, labeled in Cy3. In a single hybridization, the total input DNA serves as a baseline and is compared to the immunoprecipitated samples. Ratios of normalized signal intensities were calculated to identify enrichment of a particular sequence after immunoprecipitation, in comparison to the total input DNA. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis These data were normalized and subjected to hypothesis testing. Error rate was controlled only by Benjamini and Hochberg's step-up procedure for limiting the False Discovery Rate. WT seedlings, 9 days old This is the normalized result of the paired dye swap samples EV106 and EV107. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using false discovery rate (FDR) controlling methods. The step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found after hypothesis-testing with a controlled error rate to be significantly enriched or depleted for H3K4 methylation compared to mean values found in euchromatic regions are flagged in the FDR column. No family-wise error rate method was used to analyze this sample. Keywords: other

Project description:This is a biological replicate of EV51+52. The purpose of the chromatin immunoprecipitation/microarray (ChIP/chip) experiment is to determine which regions of a genome are enriched for a particular histone modification in a single Arabidopsis thanliana genotype. Chromatin immunoprecipitation with antibodies raised against dimethyl histone-H3 lysine-9 (H3mK9) or dimethyl histone-H3 lysine-4 (H3mK4) is performed on a selected genotype. This purified DNA from each immunoprecipiation (mH3K9, mH3K4, no antibody control) is used for random amplification to increase the quantity of DNA for microarray hybridization. The amplified DNA from each experimental sample is then labeled with Cy5 and hybridized against total input DNA from the corresponding genotype, labeled in Cy3. In a single hybridization, the total input DNA serves as a baseline and is compared to the immunoprecipitated samples. Ratios of normalized signal intensities were calculated to identify enrichment of a particular sequence after immunoprecipitation, in comparison to the total input DNA. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis These data were normalized and subjected to hypothesis testing. Error rate was controlled by Benjamini and Hochberg's step-up procedure for limiting the False Discovery Rate. Wild-type seedlings, 9 days old This is the normalized result of the paired dye swap samples EV108 and EV109. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. The style of hypothesis test proposed by Black and Doerge (2002) was applied to each of the features represented on each array, with rejection of the null hypothesis indicating a significant change in fluorescence intensity. To account for the number of hypothesis tests being made, and thus provide some level of error rate control, significance was assessed using false discovery rate (FDR) controlling methods. The step-up procedure of Benjamini and Hochberg (1995) was used to control the FDR below alpha = 0.01. For the purposes of this experiment, the hypotheses were assumed to be independent. Features found after hypothesis-testing with a controlled error rate to be significantly enriched or depleted for H3K9 methylation compared to mean values found in euchromatic regions are flagged in the column. No family-wise error rate methods were used to analyze this sample. Keywords: other

Dataset Information

EV23+24 mRNA levels in Wild-type versus ddm1/+ backcross bolting Arabidopsis thaliana plants

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