Microarray Coverage of Universal Human Reference RNA on microarrays with yeast controls
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE3973 | GEO | 2006/01/05
SECONDARY ACCESSION(S): PRJNA104877
REPOSITORIES: GEO
Similar Datasets
Project description:Microarray coverage of UHRR was determined by counting the number of spots with intensities exceeding the 1% and 5% false positive rates defined using the yeast negative control spots. Seven microarrays from different print runs were used. In four experiments, the UHRR was labeled with Cy5 and in three experiments with Cy3. Labeled UHRR was co-hybridized with Common Reference RNA (CRF) developed at Stanford University (8) labeled with the alternate dye. A reference experiment design type is where all samples are compared to a common reference. Computed
2006-01-04 | E-GEOD-3973 | biostudies-arrayexpress
Project description:Microarray coverage of UHRR was determined by counting the number of spots with intensities exceeding the 1% and 5% false positive rates defined using the yeast negative control spots. Seven microarrays from different print runs were used. In four experiments, the UHRR was labeled with Cy5 and in three experiments with Cy3. Labeled UHRR was co-hybridized with Common Reference RNA (CRF) developed at Stanford University (8) labeled with the alternate dye.
2005-11-01 | E-SMDB-2652 | biostudies-arrayexpress
Project description:Total RNA isolated from 10 individual human cell lines were reverse-transcribed to cDNA, labeled with Cy5 and co-hybridized with Cy3-labeled UHRR onto 43,000-spot cDNA microarrays (Stanford University). The data was analyzed using GeneTraffic software. Approximately 6000-8000 spots out of 43,000 (14-18 %) were flagged on each microarray and excluded from further analysis. Spots with hybridization signals in Cy5 channel higher than 1000 and with Cy5/Cy3 ratio greater than 2 were collected and the number of spots with these characteristics in only one cell line was determined. A reference experiment design type is where all samples are compared to a common reference. Keywords: reference_design
2006-01-05 | GSE3972 | GEO
Project description:RNA from 11 individual mouse cell lines were reverse-transcribed to cDNA, labeled with Cy5 and cohybridized with Cy3-labeled UMRR onto 7,500-spot mouse oligo microarrays (UNC). The data was analyzed using GeneTraffic software. 300-1000 spots out of 8,000 were flagged on each microarray and excluded from further analysis. Spots with hybridization signals in Cy5 channel higher than 1000 and with Cy5/Cy3 ratio greater than 2 were collected and the number of spots with these characteristics on only one microarray was determined. A reference experiment design type is where all samples are compared to a common reference. Keywords: reference_design
2006-01-05 | GSE3974 | GEO
Project description:Total RNA isolated from 10 individual human cell lines were reverse-transcribed to cDNA, labeled with Cy5 and co-hybridized with Cy3-labeled UHRR onto 43,000-spot cDNA microarrays (Stanford University). The data was analyzed using GeneTraffic software. Approximately 6000-8000 spots out of 43,000 (14-18 %) were flagged on each microarray and excluded from further analysis. Spots with hybridization signals in Cy5 channel higher than 1000 and with Cy5/Cy3 ratio greater than 2 were collected and the number of spots with these characteristics in only one cell line was determined.
2005-11-01 | E-SMDB-2651 | biostudies-arrayexpress
Project description:The experiment was designed to enable validation of pre-processing and test algorithms. The in-house produced cDNA-array contains 44 clones: 19 clones from mice, 4 clones from Pine (involved in photosynthesis) and 21 artificial clones from the Lucidea Universal ScoreCard (Amersham Bioscienses), where each clone was printed 480 times in 48 identically designed sub-grids. The experiment contains eight arrays with identical experimental design. Total RNA from murine cell line was divided in two samples; the reference sample was labeled with the fluorophore Cy5 and spiked with the Lucidea Universal ScoreCard reference reaction, the test sample was labeled with Cy3 and spiked with the Lucidea Universal ScoreCard test reaction. The concentrations of the Lucidea reactions are known and consequently so are the true ratios between the reference and test channels. Approximately 40% of the artificial genes were differentially expressed. The particular design of the experiment makes the data suitable for validating algorithms for pre-processing and tests for identifying differentially expressed genes. - The 80.I.1 normalization algorithm : all spots flagged as not found during image analysis considered missing values (complete filtration). Print-tip MA-loess dye normalization based on the calibration clones was applied to the raw data and the B-statistic was calculated. - The 80.II.1 normalization algorithm : all spots flagged as not found during image analysis considered missing values (complete filtration). Local background correction was applied and all spots with negative values were excluded. Print-tip MA-loess dye normalization based on the calibration clones was applied to the background corrected data and the B-statistic was calculated. - The 80.II.64 normalization algorithm : all spots flagged as not found during image analysis considered missing values (complete filtration). Local background correction was applied, negative values were excluded and all spots with a value below 64 were set to 64 (censored). Print-tip MA-loess dye normalization based on the calibration clones was applied to the background corrected data and the B-statistic was calculated. - The 80.V.1 normalization algorithm : The intensities of all spots flagged not found were treated as missing values during normalization, but prior to calculating test-statistics the spot's log-ratios were set to zero. In the special case when all arrays generated not-found spots, the gene was removed from the experiment (Partial filtration). Local background correction was applied and all negative values were considered missing values. Print-tip MA-loess dye normalization based on the calibration clones was applied to the background corrected data and the B-statistic was calculated.
2011-05-21 | GSE29400 | GEO
Project description:Abstract: BACKGROUND: Obtaining reliable and reproducible two-color microarray gene expression data is critically important for understanding the biological significance of perturbations made on a cellular system. Microarray design, RNA preparation and labeling, hybridization conditions and data acquisition and analysis are variables difficult to simultaneously control. A useful tool for monitoring and controlling intra- and inter-experimental variation is Universal Reference RNA (URR), developed with the goal of providing hybridization signal at each microarray probe location (spot). Measuring signal at each spot as the ratio of experimental RNA to reference RNA targets, rather than relying on absolute signal intensity, decreases variability by normalizing signal output in any two-color hybridization experiment. RESULTS: Human, mouse and rat URR (UHRR, UMRR and URRR, respectively) were prepared from pools of RNA derived from individual cell lines representing different tissues. A variety of microarrays were used to determine percentage of spots hybridizing with URR and producing signal above a user defined threshold (microarray coverage). Microarray coverage was consistently greater than 80% for all arrays tested. We confirmed that individual cell lines contribute their own unique set of genes to URR, arguing for a pool of RNA from several cell lines as a better configuration for URR as opposed to a single cell line source for URR. Microarray coverage comparing two separately prepared batches each of UHRR, UMRR and URRR were highly correlated (Pearson's correlation coefficients of 0.97). CONCLUSION: Results of this study demonstrate that large quantities of pooled RNA from individual cell lines are reproducibly prepared and possess diverse gene representation. This type of reference provides a standard for reducing variation in microarray experiments and allows more reliable comparison of gene expression data within and between experiments and laboratories. This SuperSeries is composed of the SubSeries listed below.
2006-03-08 | GSE4389 | GEO
Project description:This SuperSeries is composed of the following subset Series: GSE3972: Contribution of unique genes by individual cell lines to Universal Human Reference RNA GSE3973: Microarray Coverage of Universal Human Reference RNA on microarrays with yeast controls GSE3974: Contribution of unique genes by individual cell lines to Universal Mouse Reference RNA Abstract: BACKGROUND: Obtaining reliable and reproducible two-color microarray gene expression data is critically important for understanding the biological significance of perturbations made on a cellular system. Microarray design, RNA preparation and labeling, hybridization conditions and data acquisition and analysis are variables difficult to simultaneously control. A useful tool for monitoring and controlling intra- and inter-experimental variation is Universal Reference RNA (URR), developed with the goal of providing hybridization signal at each microarray probe location (spot). Measuring signal at each spot as the ratio of experimental RNA to reference RNA targets, rather than relying on absolute signal intensity, decreases variability by normalizing signal output in any two-color hybridization experiment. RESULTS: Human, mouse and rat URR (UHRR, UMRR and URRR, respectively) were prepared from pools of RNA derived from individual cell lines representing different tissues. A variety of microarrays were used to determine percentage of spots hybridizing with URR and producing signal above a user defined threshold (microarray coverage). Microarray coverage was consistently greater than 80% for all arrays tested. We confirmed that individual cell lines contribute their own unique set of genes to URR, arguing for a pool of RNA from several cell lines as a better configuration for URR as opposed to a single cell line source for URR. Microarray coverage comparing two separately prepared batches each of UHRR, UMRR and URRR were highly correlated (Pearson's correlation coefficients of 0.97). CONCLUSION: Results of this study demonstrate that large quantities of pooled RNA from individual cell lines are reproducibly prepared and possess diverse gene representation. This type of reference provides a standard for reducing variation in microarray experiments and allows more reliable comparison of gene expression data within and between experiments and laboratories. Refer to individual Series
2006-03-07 | E-GEOD-4389 | biostudies-arrayexpress
Project description:Many library preparation methods are available for gene expression quantification. Here, we sequenced and analysed Universal Human Reference RNA (UHRR) prepared using Smart-Seq2, TruSeq (public data) and a protocol using unique molecular identifiers (UMIs) that all include the ERCC spike-in mRNAs to investigate the effects of amplification bias on expression quantification.
2016-05-13 | GSE75823 | GEO
Project description:RNA from 11 individual mouse cell lines were reverse-transcribed to cDNA, labeled with Cy5 and cohybridized with Cy3-labeled UMRR onto 7,500-spot mouse oligo microarrays (UNC). The data was analyzed using GeneTraffic software. 300-1000 spots out of 8,000 were flagged on each microarray and excluded from further analysis. Spots with hybridization signals in Cy5 channel higher than 1000 and with Cy5/Cy3 ratio greater than 2 were collected and the number of spots with these characteristics on only one microarray was determined. A reference experiment design type is where all samples are compared to a common reference. Computed
2006-01-04 | E-GEOD-3974 | biostudies-arrayexpress