A systematic spatial bias in microarray hybridizations caused by probe spot position-dependent variability in lateral diffusion
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA135141 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay format for high-throughput detection of many parallel targetsin medical and biological research. Though commonly applied, microarray technology still suffers limitations arising from problems of data robustness and reproducibility across platforms, stemming in part from an incomplete understanding of the complex processes governing surface hybridization behavior. It has been observed that there are non-random spatial variations within individual microarray hybridizations, but the causative mechanisms of positional bias remain largely unexplained. This study identifies a symptomatic spatial bias in surface hybridization signal intensitywith systematically increased signal intensities of spots located at the boundaries of the spotted areas of the microarray slide and characterizes the underlying mechanistic principle of this bias using a simplified block array format. Experimentally-derived hybridization dynamics are compared with a mathematical modeling analysis, which together showthat the driver of the spatial bias is a position-dependent variation in lateral diffusion. Numerical simulations employing a diffusion-based model are used to demonstrate the strong influence of microarray well geometry on this spatial bias and to determine optimal conditions for which this bias can be minimized or eliminated, resulting in increased uniformity of microarray hybridization. A simplified square array of 900 identical spotted probes was hybridized using different target concentrations and different time periods to analyze systematically spatial biases in microarray hybridizations.
2010-12-22 | E-GEOD-26275 | biostudies-arrayexpress
Project description:The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay format for high-throughput detection of many parallel targetsin medical and biological research. Though commonly applied, microarray technology still suffers limitations arising from problems of data robustness and reproducibility across platforms, stemming in part from an incomplete understanding of the complex processes governing surface hybridization behavior. It has been observed that there are non-random spatial variations within individual microarray hybridizations, but the causative mechanisms of positional bias remain largely unexplained. This study identifies a symptomatic spatial bias in surface hybridization signal intensitywith systematically increased signal intensities of spots located at the boundaries of the spotted areas of the microarray slide and characterizes the underlying mechanistic principle of this bias using a simplified block array format. Experimentally-derived hybridization dynamics are compared with a mathematical modeling analysis, which together showthat the driver of the spatial bias is a position-dependent variation in lateral diffusion. Numerical simulations employing a diffusion-based model are used to demonstrate the strong influence of microarray well geometry on this spatial bias and to determine optimal conditions for which this bias can be minimized or eliminated, resulting in increased uniformity of microarray hybridization.
2010-12-22 | GSE26275 | GEO
Project description:BackgroundThe hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay for the parallel detection of multiple targets in medical and biological research. Despite its widespread application, DNA microarray technology still suffers from several biases and lack of reproducibility, stemming in part from an incomplete understanding of the processes governing surface hybridization. In particular, non-random spatial variations within individual microarray hybridizations are often observed, but the mechanisms underpinning this positional bias remain incompletely explained.Methodology/principal findingsThis study identifies and rationalizes a systematic spatial bias in the intensity of surface hybridization, characterized by markedly increased signal intensity of spots located at the boundaries of the spotted areas of the microarray slide. Combining observations from a simplified single-probe block array format with predictions from a mathematical model, the mechanism responsible for this bias is found to be a position-dependent variation in lateral diffusion of target molecules. Numerical simulations reveal a strong influence of microarray well geometry on the spatial bias.ConclusionsReciprocal adjustment of the size of the microarray hybridization chamber to the area of surface-bound probes is a simple and effective measure to minimize or eliminate the diffusion-based bias, resulting in increased uniformity and accuracy of quantitative DNA microarray hybridization.
| S-EPMC3157431 | biostudies-literature
Project description:microRNA-seq in circCDYL probe and NC probe
| PRJNA730505 | ENA
Project description:Microfluidic devices provide a low-input and efficient platform for single-cell RNA-seq (scRNA-Seq). Here we present microfluidic diffusion-based RNA-seq (MID-RNA-seq) for conducting scRNA-seq with a diffusion-based reagent swapping scheme. This device incorporates cell trapping, lysis, reverse transcription and PCR amplification all in one microfluidic chamber. MID-RNA-Seq provides high data quality that is comparable to existing scRNA-seq methods while implementing a simple device design that permits multiplexing. The robustness and scalability of MID-RNA-Seq device will be important for transcriptomic studies of scarce cell samples.
2019-02-25 | GSE119271 | GEO
Project description:Classic ‘position effect’ experiments repositioned genes to the telomere to demonstrate that the epigenetic landscape can dramatically alter gene expression. Here we show that systematic gene knockout collections provide an exceptional resource for interrogating position effects, not only at the telomere but at every single genetic locus. Because deleted genes are replaced by the same reporter gene, interrogation of this reporter provides a sensitive probe into many different chromatin environments while controlling for genetic context. Using this approach we find that, whereas replacement of yeast genes with the kanMX marker does not perturb the chromatin landscape, differences due to gene position account for more than 35% of marker gene activity. We observe chromatin influences different from those reported previously, including an antagonistic interaction between histone H3 lysine 36 trimethylation (H3K36me3) and the Rap1 transcriptional activation site in kanMX that is mediated through a Set2-Rpd3-dependent pathway. This interaction explains why some yeast genes have been resistant to deletion and allows successful generation of these deletion strains using a modified transformation procedure. These findings demonstrate that chromatin regulation is not governed by a uniform ‘histone code’, but by specific interactions between chromatin and genetic factors.
2012-11-28 | GSE42554 | GEO
Project description:RNASeq of roots from two genotypes of Arabidopsis thaliana plants, Col-0 and myb36-2 grown axenically or with a 41 member bacterial Synthetic Community (SynCom) to explore the interaction between the root diffusion barriers and the root microbiome.
2020-11-01 | GSE151376 | GEO
Project description:Modeling Formamide Denaturation of Probe-Target Hybrids for Improved Microarray Probe Design in Microbial Diagnostics
| PRJNA148019 | ENA
Project description:Classic ‘position effect’ experiments repositioned genes to the telomere to demonstrate that the epigenetic landscape can dramatically alter gene expression. Here we show that systematic gene knockout collections provide an exceptional resource for interrogating position effects, not only at the telomere but at every single genetic locus. Because deleted genes are replaced by the same reporter gene, interrogation of this reporter provides a sensitive probe into many different chromatin environments while controlling for genetic context. Using this approach we find that, whereas replacement of yeast genes with the kanMX marker does not perturb the chromatin landscape, differences due to gene position account for more than 35% of marker gene activity. We observe chromatin influences different from those reported previously, including an antagonistic interaction between histone H3 lysine 36 trimethylation (H3K36me3) and the Rap1 transcriptional activation site in kanMX that is mediated through a Set2-Rpd3-dependent pathway. This interaction explains why some yeast genes have been resistant to deletion and allows successful generation of these deletion strains using a modified transformation procedure. These findings demonstrate that chromatin regulation is not governed by a uniform ‘histone code’, but by specific interactions between chromatin and genetic factors. Data included are RNA-Seq data for 4 heterzygous diploid yeast strains and diploid wild-type. Therea re three replicates for each heterzygous strain, and six replicates for wild-type.
2012-11-28 | E-GEOD-42554 | biostudies-arrayexpress