A reanalysis of a published Affymetrix GeneChip control dataset.
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ABSTRACT:
SUBMITTER: Dabney AR
PROVIDER: S-EPMC1557755 | biostudies-literature | 2006
REPOSITORIES: biostudies-literature
Publications
A reanalysis of a published Affymetrix GeneChip control dataset.
Dabney Alan R AR Storey John D JD
Genome biology 20060322 3
A response to Preferred analysis methods for Affymetrix GeneChips revealed by a wholly defined control dataset by SE Choe, M Boutros, AM Michelson, GM Church and MS Halfon. Genome Biology 2005, 6:R16. ...[more]
Similar Datasets
Project description:A comment on Preferred analysis methods for Affymetrix GeneChips revealed by a wholly defined control dataset by SE Choe, M Boutros, AM Michelson, GM Church and MS Halfon. Genome Biology 2005, 6:R16.
| S-EPMC1779590 | biostudies-literature
Project description:BackgroundMicroarray technology has become a widely used tool in the biological sciences. Over the past decade, the number of users has grown exponentially, and with the number of applications and secondary data analyses rapidly increasing, we expect this rate to continue. Various initiatives such as the External RNA Control Consortium (ERCC) and the MicroArray Quality Control (MAQC) project have explored ways to provide standards for the technology. For microarrays to become generally accepted as a reliable technology, statistical methods for assessing quality will be an indispensable component; however, there remains a lack of consensus in both defining and measuring microarray quality.ResultsWe begin by providing a precise definition of microarray quality and reviewing existing Affymetrix GeneChip quality metrics in light of this definition. We show that the best-performing metrics require multiple arrays to be assessed simultaneously. While such multi-array quality metrics are adequate for bench science, as microarrays begin to be used in clinical settings, single-array quality metrics will be indispensable. To this end, we define a single-array version of one of the best multi-array quality metrics and show that this metric performs as well as the best multi-array metrics. We then use this new quality metric to assess the quality of microarry data available via the Gene Expression Omnibus (GEO) using more than 22,000 Affymetrix HGU133a and HGU133plus2 arrays from 809 studies.ConclusionsWe find that approximately 10 percent of these publicly available arrays are of poor quality. Moreover, the quality of microarray measurements varies greatly from hybridization to hybridization, study to study, and lab to lab, with some experiments producing unusable data. Many of the concepts described here are applicable to other high-throughput technologies.
| S-EPMC3097162 | biostudies-literature
Project description:Since the introduction of microarrays in 1995, researchers world-wide have used both commercial and custom-designed microarrays for understanding differential expression of transcribed genes. Public databases such as ArrayExpress and the Gene Expression Omnibus (GEO) have made millions of samples readily available. One main drawback to microarray data analysis involves the selection of probes to represent a specific transcript of interest, particularly in light of the fact that transcript-specific knowledge (notably alternative splicing) is dynamic in nature.We therefore developed a framework for reannotating and reassigning probe groups for Affymetrix® GeneChip® technology based on functional regions of interest. This framework addresses three issues of Affymetrix® GeneChip® data analyses: removing nonspecific probes, updating probe target mapping based on the latest genome knowledge and grouping probes into gene, transcript and region-based (UTR, individual exon, CDS) probe sets. Updated gene and transcript probe sets provide more specific analysis results based on current genomic and transcriptomic knowledge. The framework selects unique probes, aligns them to gene annotations and generates a custom Chip Description File (CDF). The analysis reveals only 87% of the Affymetrix® GeneChip® HG-U133 Plus 2 probes uniquely align to the current hg38 human assembly without mismatches. We also tested new mappings on the publicly available data series using rat and human data from GSE48611 and GSE72551 obtained from GEO, and illustrate that functional grouping allows for the subtle detection of regions of interest likely to have phenotypical consequences.Through reanalysis of the publicly available data series GSE48611 and GSE72551, we profiled the contribution of UTR and CDS regions to the gene expression levels globally. The comparison between region and gene based results indicated that the detected expressed genes by gene-based and region-based CDFs show high consistency and regions based results allows us to detection of changes in transcript formation.
| S-EPMC5731501 | biostudies-literature
Project description:Affymetrix GeneChip microarrays are the most widely used high-throughput technology to measure gene expression, and a wide variety of preprocessing methods have been developed to transform probe intensities reported by a microarray scanner into gene expression estimates. There have been numerous comparisons of these preprocessing methods, focusing on the most common analyses-detection of differential expression and gene or sample clustering. Recently, more complex multivariate analyses, such as gene co-expression, differential co-expression, gene set analysis and network modeling, are becoming more common; however, the same preprocessing methods are typically applied. In this article, we examine the effect of preprocessing methods on some of these multivariate analyses and provide guidance to the user as to which methods are most appropriate.
| S-EPMC3431718 | biostudies-literature
Project description:The experimental spike-in studies of microarray hybridization conducted by Affymetrix demonstrate a nonlinear response of fluorescence intensity signal to target concentration. Several theoretical models have been put forward to explain these data. It was shown that the Langmuir adsorption isotherm recapitulates a general trend of signal response to concentration. However, this model fails to explain some key properties of the observed signal. In particular, according to the simple Langmuir isotherm, all probes should saturate at the same intensity level. However, this effect was not observed in the publicly available Affymetrix spike-in data sets. On the contrary, it was found that the saturation intensities vary greatly and can be predicted based on the probe sequence composition. In our experimental study, we attempt to account for the unexplained variation in the observed probe intensities using customized fluidics scripts. We explore experimentally the effect of the stringent wash, target concentration and hybridization time on the final microarray signal. The washing effect is assessed by scanning chips both prior to and after the stringent wash. Selective labeling of both specific and non-specific targets allows the visualization and investigation of the washing effect for both specific and non-specific signal components. We propose a new qualitative model of the probe-target hybridization mechanism that is in agreement with observed hybridization and washing properties of short oligonucleotide microarrays. This study demonstrates that desorption of incompletely bound targets during the washing cycle contributes to the observed difference in saturation levels.
| S-EPMC1919478 | biostudies-literature
Project description:BackgroundHigh throughput microarray-based single nucleotide polymorphism (SNP) genotyping has revolutionized the way genome-wide linkage scans and association analyses are performed. One of the key features of the array-based GeneChip Mapping 10K Array from Affymetrix is the automated SNP calling algorithm. The Affymetrix algorithm was trained on a database of ethnically diverse DNA samples to create SNP call zones that are used as static models to make genotype calls for experimental data. We describe here the implementation of clustering algorithms on large training datasets resulting in improved SNP call rates on the 10K GeneChip.ResultsA database of 948 individuals genotyped on the GeneChip Mapping 10K 2.0 Array was used to identify 822 SNPs that were called consistently less than 75% of the time. These SNPs represent on average 8.25% of the total SNPs on each chromosome with chromosome 19, the most gene-rich chromosome, containing the highest proportion of poor performers (18.7%). To remedy this, we created SNiPer, a new application which uses two clustering algorithms to yield increased call rates and equivalent concordance to Affymetrix called genotypes. We include a training set for these algorithms based on individual genotypes for 705 samples. SNiPer has the capability to be retrained for lab-specific training sets. SNiPer is freely available for download at http://www.tgen.org/neurogenomics/data.ConclusionThe correct calling of poor performing SNPs may prove to be key in future linkage studies performed on the 10K GeneChip. It would prove particularly invaluable for those diseases that map to chromosome 19, known to contain a high proportion of poorly performing SNPs. Our results illustrate that SNiPer can be used to increase call rates on the 10K GeneChip without sacrificing accuracy, thereby increasing the amount of valid data generated.
| S-EPMC1280925 | biostudies-literature
Project description:BACKGROUND: Affymetrix GeneChip typically contains multiple probe sets per gene, defined as sibling probe sets in this study. These probe sets may or may not behave similar across treatments. The most appropriate way of consolidating sibling probe sets suitable for analysis is an open problem. We propose the Analysis of Variance (ANOVA) framework to decide which sibling probe sets can be consolidated. RESULTS: The ANOVA model allows us to separate the sibling probe sets into two types: those behave similarly across treatments and those behave differently across treatments. We found that consolidation of sibling probe sets of the former type results in large increase in the number of differentially expressed genes under various statistical criteria. The approach to selecting sibling probe sets suitable for consolidating is implemented in R language and freely available from http://research.stowers-institute.org/hul/affy/. CONCLUSION: Our ANOVA analysis of sibling probe sets provides a statistical framework for selecting sibling probe sets for consolidation. Consolidating sibling probe sets by pooling data from each greatly improves the estimates of a gene expression level and results in identification of more biologically relevant genes. Sibling probe sets that do not qualify for consolidation may represent annotation errors or other artifacts, or may correspond to differentially processed transcripts of the same gene that require further analysis.
| S-EPMC2397416 | biostudies-literature
Project description:Affymetrix exon arrays offer scientists the only solution for exon-level expression profiling at the whole-genome scale on a single array. These arrays feature a new chip design with no mismatch probes and a radically new random primed protocol to generate sense DNA targets along the entire length of the transcript. In addition to these changes, a limited number of validating experiments and virtually no experimental data to rigorously address the comparability of all-exon arrays with conventional 3'-arrays result in a natural reluctance to replace conventional expression arrays with the new all-exon platform.Using commercially available Affymetrix arrays, we assess the performance of the Human Exon 1.0 ST (HuEx) and U133 Plus 2.0 (U133Plus2) platforms directly through a series of 'spike-in' hybridizations containing 25 transcripts in the presence of a fixed eukaryotic background. Specifically, we compare the measures of expression for HuEx and U133Plus2 arrays to evaluate the precision of these measures as well as the specificity and sensitivity of the measures' ability to detect differential expression.This study presents an experimental comparison and systematic cross-validation of Affymetrix exon arrays and establishes high comparability of expression changes and probe performance characteristics between Affymetrix conventional and exon arrays. In addition, this study offers a reliable benchmark data set for the comparison of competing exon expression measures, the selection of methods suitable for mapping exon array measures to the wealth of previously generated microarray data, as well as the development of more advanced methods for exon- and transcript-level expression summarization.
| S-EPMC1976394 | biostudies-literature
Project description:Improvements in technology have made it possible to conduct genome-wide association mapping at costs within reach of academic investigators, and experiments are currently being conducted with a variety of high-throughput platforms. To provide an appropriate context for interpreting results of such studies, we summarize here results of an investigation of one of the first of these technologies to be publicly available, the Affymetrix GeneChip Human Mapping 100K set of single nucleotide polymorphisms (SNPs). In a systematic analysis of the pattern and distribution of SNPs in the Mapping 100K set, we find that SNPs in this set are undersampled from coding regions (both nonsynonymous and synonymous) and oversampled from regions outside genes, relative to SNPs in the overall HapMap database. In addition, we utilize a novel multilocus linkage disequilibrium (LD) coefficient based on information content (analogous to the information content scores commonly used for linkage mapping) that is equivalent to the familiar measure r2 in the special case of two loci. Using this approach, we are able to summarize for any subset of markers, such as the Affymetrix Mapping 100K set, the information available for association mapping in that subset, relative to the information available in the full set of markers included in the HapMap, and highlight circumstances in which this multilocus measure of LD provides substantial additional insight about the haplotype structure in a region over pairwise measures of LD.
| S-EPMC1456318 | biostudies-literature
Project description:BackgroundAs more methods are developed to analyze RNA-profiling data, assessing their performance using control datasets becomes increasingly important.ResultsWe present a 'spike-in' experiment for Affymetrix GeneChips that provides a defined dataset of 3,860 RNA species, which we use to evaluate analysis options for identifying differentially expressed genes. The experimental design incorporates two novel features. First, to obtain accurate estimates of false-positive and false-negative rates, 100-200 RNAs are spiked in at each fold-change level of interest, ranging from 1.2 to 4-fold. Second, instead of using an uncharacterized background RNA sample, a set of 2,551 RNA species is used as the constant (1x) set, allowing us to know whether any given probe set is truly present or absent. Application of a large number of analysis methods to this dataset reveals clear variation in their ability to identify differentially expressed genes. False-negative and false-positive rates are minimized when the following options are chosen: subtracting nonspecific signal from the PM probe intensities; performing an intensity-dependent normalization at the probe set level; and incorporating a signal intensity-dependent standard deviation in the test statistic.ConclusionsA best-route combination of analysis methods is presented that allows detection of approximately 70% of true positives before reaching a 10% false-discovery rate. We highlight areas in need of improvement, including better estimate of false-discovery rates and decreased false-negative rates.
| S-EPMC551536 | biostudies-literature