IQRray, a new method for Affymetrix microarray quality control, and the homologous organ conservation score, a new benchmark method for quality control metrics

Motivation: Microarray results accumulated in public repositories are widely reused in meta-analytical studies and secondary databases. The quality of the data obtained with this technology varies from experiment to experiment, and an efficient method for quality assessment is necessary to ensure their reliability. Results: The lack of a good benchmark has hampered evaluation of existing methods for quality control. In this study, we propose a new independent quality metric that is based on evolutionary conservation of expression profiles. We show, using 11 large organ-specific datasets, that IQRray, a new quality metrics developed by us, exhibits the highest correlation with this reference metric, among 14 metrics tested. IQRray outperforms other methods in identification of poor quality arrays in datasets composed of arrays from many independent experiments. In contrast, the performance of methods designed for detecting outliers in a single experiment like Normalized Unscaled Standard Error and Relative Log Expression was low because of the inability of these methods to detect datasets containing only low-quality arrays and because the scores cannot be directly compared between experiments. Availability and implementation: The R implementation of IQRray is available at: ftp://lausanne.isb-sib.ch/pub/databases/Bgee/general/IQRray.R. Contact: [email protected] Supplementary information: Supplementary data are available at Bioinformatics onlin

Blood lactose after dairy product intake in healthy men.

The absence of a dedicated transport for disaccharides in the intestine implicates that the metabolic use of dietary lactose relies on its prior hydrolysis at the intestinal brush border. Consequently, lactose in blood or urine has mostly been associated with specific cases in which the gastrointestinal barrier is damaged. On the other hand, lactose appears in the blood of lactating women and has been detected in the blood and urine of healthy men, indicating that the presence of lactose in the circulation of healthy subjects is not incompatible with normal physiology. In this cross-over study we have characterised the postprandial kinetics of lactose, and its major constituent, galactose, in the serum of fourteen healthy men who consumed a unique dose of 800 g milk or yogurt. Genetic testing for lactase persistence and microbiota profiling of the subjects were also performed. Data revealed that lactose does appear in serum after dairy intake, although with delayed kinetics compared with galactose. Median serum concentrations of approximately 0·02 mmol/l lactose and approximately 0·2 mmol/l galactose were observed after the ingestion of milk and yogurt respectively. The serum concentrations of lactose were inversely correlated with the concentrations of galactose, and the variability observed between the subjects' responses could not be explained by the presence of the lactase persistence allele. Finally, lactose levels have been associated with the abundance of the Veillonella genus in faecal microbiota. The measurement of systemic lactose following dietary intake could provide information about lactose metabolism and nutrient transport processes under normal or pathological conditions

IQRray, a new method for Affymetrix microarray quality control, and the homologous organ conservation score, a new benchmark method for quality control metrics.

MOTIVATION: Microarray results accumulated in public repositories are widely reused in meta-analytical studies and secondary databases. The quality of the data obtained with this technology varies from experiment to experiment, and an efficient method for quality assessment is necessary to ensure their reliability. RESULTS: The lack of a good benchmark has hampered evaluation of existing methods for quality control. In this study, we propose a new independent quality metric that is based on evolutionary conservation of expression profiles. We show, using 11 large organ-specific datasets, that IQRray, a new quality metrics developed by us, exhibits the highest correlation with this reference metric, among 14 metrics tested. IQRray outperforms other methods in identification of poor quality arrays in datasets composed of arrays from many independent experiments. In contrast, the performance of methods designed for detecting outliers in a single experiment like Normalized Unscaled Standard Error and Relative Log Expression was low because of the inability of these methods to detect datasets containing only low-quality arrays and because the scores cannot be directly compared between experiments. AVAILABILITY AND IMPLEMENTATION: The R implementation of IQRray is available at: ftp://lausanne.isb-sib.ch/pub/databases/Bgee/general/IQRray.R. CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online

Microarray transciptome analysis of Arabidopsis thaliana mutant abh1

Wydział Biologii: Instytut Biologii Molekularnej i BiotechnologiiMutant abh1 Arabidopsis thaliana posiadający insercję T-DNA w genie kodującym mniejszą podjednostkę jądrowego kompleksu wiążącego się z kapem (CBC) wykazuje nadwrażliwość na kwas abscysynowy (ABA) podczas kiełkowania nasion. W celu poznania zmian towarzyszących odmiennej odpowiedzi tego mutanta na ABA w niskim stężeniu przeprowadziliśmy analizę mikromacierzową z użyciem mikromacierzy tilingowych transkryptomu nasion mutanta abh1 i roślin typu dzikiego kiełkujących w obecności ABA i bez. Uzyskane przez nas wyniki wskazują, że w nasionach mutanta pod wpływem ABA dochodzi do silnej aktywacji transpozonów. Analizę genów ulegających różnicowej ekspresji przeprowadzono za pomocą nowego algorytmu opartego na pozycjach Difference of Rank Products (DRP), który jak udowodniłam najlepiej nadaje się do analizy eksperymentów mikromacierzowych z niewielką liczbą powtórzeń. W celu poznania genów, które współuczestniczą z kompleksem CBC w przekazywaniu sygnału ABA przeprowadziliśmy mutagenezę chemiczną mutanta abh1, dzięki której udało nam się uzyskać 5 linii supresorowych o zniesionej nadwrażliwości nasion na ABA. Mutacja w linii A21 została zmapowana do dolnego ramienia chromosomu II, co doprowadziło nas do odkrycia mutacji punktowej w genie ABI5. Sekwencjonowanie genów ABI4 i ABI5 ujawniło istnienie mutacji nonsensowych w genie ABI4 w dwóch innych liniach supresorowych.The nuclear cap binding complex (CBC) consists of two subunits: CBP20 and CBP80. Arabidopsis thaliana mutant abh1 with T-DNA insertion in the CBP80 gene is hypersensitive to plant hormone abscisic acid (ABA) during seed germination. To investigate transcriptomic changes that correspond to response of abh1 mutant to ABA, we performed tiling microarray experiment on germinating seeds of the abh1 and wild type plants treated and untreated with low concentration of abscisic acid. The obtained results indicate that during germination of abh1 seeds in the presence of abscisic acid occur massive activation of transposons. The analysis of differentially expressed genes were carried out with newly proposed rank-based algorithm Difference of Rank Products (DRP), which we proved is the best for analysis of microarray experiments with low number of samples. To identify genes that cooperate with CBC in ABA signal transmission we performed chemical mutagenesis of the abh1 mutants. Among progeny of EMS treated plants we selected five independent suppressors of exaggerated reaction of abh1 seeds to of abscisic acid. Mutation in the line A21 was mapped to the bottom arm of chromosome 2 which lead us to identification of a point mutation in ABI5 gene. Sequencing of the ABI4 and ABI5 from other suppressor lines revealed existence of nonsense mutations in the ABI4 gene in two other lines

Microarray transciptome analysis of Arabidopsis thaliana mutant abh1

Wydział Biologii: Instytut Biologii Molekularnej i BiotechnologiiMutant abh1 Arabidopsis thaliana posiadający insercję T-DNA w genie kodującym mniejszą podjednostkę jądrowego kompleksu wiążącego się z kapem (CBC) wykazuje nadwrażliwość na kwas abscysynowy (ABA) podczas kiełkowania nasion. W celu poznania zmian towarzyszących odmiennej odpowiedzi tego mutanta na ABA w niskim stężeniu przeprowadziliśmy analizę mikromacierzową z użyciem mikromacierzy tilingowych transkryptomu nasion mutanta abh1 i roślin typu dzikiego kiełkujących w obecności ABA i bez. Uzyskane przez nas wyniki wskazują, że w nasionach mutanta pod wpływem ABA dochodzi do silnej aktywacji transpozonów. Analizę genów ulegających różnicowej ekspresji przeprowadzono za pomocą nowego algorytmu opartego na pozycjach Difference of Rank Products (DRP), który jak udowodniłam najlepiej nadaje się do analizy eksperymentów mikromacierzowych z niewielką liczbą powtórzeń. W celu poznania genów, które współuczestniczą z kompleksem CBC w przekazywaniu sygnału ABA przeprowadziliśmy mutagenezę chemiczną mutanta abh1, dzięki której udało nam się uzyskać 5 linii supresorowych o zniesionej nadwrażliwości nasion na ABA. Mutacja w linii A21 została zmapowana do dolnego ramienia chromosomu II, co doprowadziło nas do odkrycia mutacji punktowej w genie ABI5. Sekwencjonowanie genów ABI4 i ABI5 ujawniło istnienie mutacji nonsensowych w genie ABI4 w dwóch innych liniach supresorowych.The nuclear cap binding complex (CBC) consists of two subunits: CBP20 and CBP80. Arabidopsis thaliana mutant abh1 with T-DNA insertion in the CBP80 gene is hypersensitive to plant hormone abscisic acid (ABA) during seed germination. To investigate transcriptomic changes that correspond to response of abh1 mutant to ABA, we performed tiling microarray experiment on germinating seeds of the abh1 and wild type plants treated and untreated with low concentration of abscisic acid. The obtained results indicate that during germination of abh1 seeds in the presence of abscisic acid occur massive activation of transposons. The analysis of differentially expressed genes were carried out with newly proposed rank-based algorithm Difference of Rank Products (DRP), which we proved is the best for analysis of microarray experiments with low number of samples. To identify genes that cooperate with CBC in ABA signal transmission we performed chemical mutagenesis of the abh1 mutants. Among progeny of EMS treated plants we selected five independent suppressors of exaggerated reaction of abh1 seeds to of abscisic acid. Mutation in the line A21 was mapped to the bottom arm of chromosome 2 which lead us to identification of a point mutation in ABI5 gene. Sequencing of the ABI4 and ABI5 from other suppressor lines revealed existence of nonsense mutations in the ABI4 gene in two other lines

Robust inference of expression state in bulk and single-cell RNA-Seq using curated intergenic regions

RNA-Seq is a powerful technique to provide quantitative information on gene expression. While many applications focus on estimated expression levels, it is also important to determine which genes are actively transcribed, and which are not. The problem can be viewed as simply setting a biologically meaningful threshold for calling a gene expressed. We propose to define this threshold per sample relative to the background level for non-expressed genomic features, inferred by the amount of reads mapped to intergenic regions of the genome. To this aim, we first define a stringent set of reference intergenic regions, based on available bulk RNA-Seq libraries for each species. We provide predefined regions selected for different animal species with varying genome annotation quality through the Bgee database. We then call genes expressed if their level of expression is significantly higher than the background noise. This approach can be applied to bulk as well as single-cell RNA-Seq, on a single library as well as on a combination of libraries over one condition. We show that the estimated proportion of expressed genes is biologically meaningful and stable between libraries originating from the same tissue, in both model and non-model organisms.Competing Interest StatementThe authors have declared no competing interest

Uncovering hidden duplicated content in public transcriptomics data.

As part of the development of the database Bgee (a dataBase for Gene Expression Evolution), we annotate and analyse expression data from different types and different sources, notably Affymetrix data from GEO and ArrayExpress, and RNA-Seq data from SRA. During our quality control procedure, we have identified duplicated content in GEO and ArrayExpress, affecting ∼14% of our data: fully or partially duplicated experiments from independent data submissions, Affymetrix chips reused in several experiments, or reused within an experiment. We present here the procedure that we have established to filter such duplicates from Affymetrix data, and our procedure to identify future potential duplicates in RNA-Seq data. Database URL: http://bgee.unil.ch

POSTPRANDIAL SERUM LACTOSE AFTER ACUTE INTAKE OF MILK AND YOGHURT

The use of dietary lactose as a source of energy requires its prior hydrolysis at the intestinal brush border into glucose and galactose. Indeed, as a disaccharide, intact lactose cannot be actively absorbed. Therefore, the presence of unhydrolysed lactose in plasma or urine has mostly been associated with specific cases such as lactating women or altered gastrointestinal permeability. Nevertheless, lactose has also been detected in the blood and urine of healthy men, indicating that the presence of lactose in the circulation of healthy subjects is not incompatible with normal physiology. The present crossover study monitored, in 14 healthy men, the postprandial appearance of lactose in serum after a single intake of 800 g of milk or yoghurt. Genetic testing for lactase persistence and microbiota profiling of the subjects were also performed in order to investigate their potential contribution to postprandial lactose serum levels. Data revealed that lactose does appear in serum after dairy intake, and with delayed kinetics when compared to the actively absorbed galactose. A notable inter-individual variability was observed and, although lactose levels were inversely correlated with galactose levels, the presence of the lactase persistence allele could not explain this variability. Finally, lactose levels have been associated with the abundance of the Veillonella genus in faecal microbiota. The measurement of systemic lactose following dietary intake could provide information about lactose metabolism and nutrient transport processes under normal or pathological conditions