Introduction to Microarray Analysis - Stratagene ArrayAssist

Transcriptomics Patrick Kemmeren European Bioinformatics Institute Genomics Lab, UMC Utrecht mRNA cDNA hybridise to microarray What are microarrays ? Transcriptomics? hybridisation labelled nucleic acid array RNA extract Sample Array design hybridisation labelled nucleic acid array RNA extract Sample hybridisation labelled nucleic acid array RNA extract Sample hybridisation labelled nucleic acid array RNA extract Sample hybridisation labelled nucleic acid Microarray RNA extract Sample Experiment Gene expression data matrix normalization integration Protocol Protocol Protocol Protocol Protocol Protocol genes Samples Genes Gene expression levels Sample annotation Gene annotation Gene expression matrix Microarray data and annotation Traditions of data sharing in Life Sciences • Data used in publications should be made available so that • the experiments can be reproduced and the conclusions can be verified • the others can build on other’s results • In genome sequencing this has evolved into submissions to public sequence databases DDBJ/EMBL/Genbank – most journals require such submissions Array scans Spots Quantitations Genes Samples A B C D Sharing microarray data – which data? MGED standards - MIAME ArraySample • Sample source • Sample treatments • Extraction protocol • Labeling protocol • Array design information • Location of each element • Description of each element Hybridization protocol • Quantification matrix • Analysis protocol • Software specifications • Image • Scanning protocol • Software specifications Hybridisation MIAME 6 parts of a microarray experiment MGED – MIAME Microarray experiment Samples Extracts Labelled Extracts Colours related to labels Hybridizations Shapes related to array designs Experiment name Rustici et al., S. pombe cell-cycle mutant data (2004) MIAMExpress Database Submissions Database Retrieval of raw & processed data for analysis Gene, sample, and experiment centric queries, Submission support Curation MAGE- ML XML VisualisationData download Data upload User Functionality Database Architecture MAGE- ML External Application ArrayExpress Repository AE Data Warehouse [...]... output Extracts 1…n Extracts 1…n Clone Tracker Labels 1…n Labels 1…n Protocol Submission Sample protocol Extraction protocol Labelling protocol Hybridisations Hyb protocol ADF generation Array1 Array2 Arrayn Array Details Data1 Data2 Datan Combined Experiment Data Submit Submit Scanning protocol Image analysis protocol Transformation protocol Submit MAGE-ML export using MAGEstk API ArrayExpress Curation... suite of tools • Standalone tool • Locally installed (UJI, UMC Utrecht) • Teaching tool • Pipelines, workflows, high-throughput analysis Acknowledgements EBI Microarray Informatics Team Alvis Brazma, Head of Microarray Informatics Group Ahmet Oezcimen, Scientist (Oracle DBA) Anastasia Samsonova, PhD student Anjan Sharma, Scientist (Software Developer) Anna Farne, Scientist (Curation) Aurora Torrente,... New! http://www.ebi.ac.uk/aedw/ArrayExpress_main.html Gene-centric Query Prototype New! Gene-centric Query Prototype - Driven by a BioMart backend New! Gene-centric Query Prototype External Application MAGEML Submission support Curation Database Architecture MAGEML XML AE Data Warehouse MIAMExpress Database ArrayExpress Repository Data upload Data download Visualisation User Submissions Database Retrieval...MIAMExpress • Submission and annotation tool • Potential local data annotation tool • Based on MIAME concepts • Accepts protocol, array and experiment submissions • User accounts allow re-use of protocols and arrays • Works with your own or commercial arrays MIAMExpress schema Create Account Array Submission File Merging... Durinck, Student Susanna-Assunta Sansone, Scientist (Toxicogenomics Proj Coordinator) Tim Rayner, Scientist (Curation) Ugis Sarkans, Scientist (Database Development Coordinator) Original EP Development: • Jaak Vilo (Tartu) • Patrick Kemmeren (Utrecht) • Misha Kapushesky EP:NG Framework Development: • Patrick Kemmeren (Utrecht) • Misha Kapushesky • Caroline Johnston (UCL) Visualization Components: • Misha Kapushesky... Architecture MAGEML XML AE Data Warehouse MIAMExpress Database ArrayExpress Repository Data upload Data download Visualisation User Submissions Database Retrieval of raw & processed data for analysis Functionality Gene, sample, and experiment centric queries, ArrayExpress http://www.ebi.ac.uk/arrayexpress • A public repository for microarray data at the EBI Data in ArrayExpress Hybridizations 8000 7000 6000... Misha Kapushesky • Steffen Durinck (Leuven) • Phil Hyoun Lee Clustering Comparison: • Aurora Torrente • Christine Körner (Leipzig) PCA/COA/BGA: • Aedín Culhane (Cork) Signature Algorithm: • Jan Ihmels (Tel-Aviv) Gene Ordering: • Karlis Freivalds (Riga) Normalisation: • Caroline Johnston (UCL) Web Services: • Antonio Estruch (UJI) ... Coordinator) Jaak Vilo, Consultant Lev Soinov, Scientist (Postdoc Wellcome Trust) Misha Kapushesky, Scientist (Scientific Application Programmer) Mohammadreza Shojatalab, Scientist (Database Programmer) Niran Abeygunawardena, Scientist (Web Designer/Programmer) Patrick Kemmeren, Consultant Per Lilja, Scientist (Database Programmer) Philippe Rocca-Serra, Scientist (Nutrigenomics Proj Coordinator) Pierre... http://www.ebi.ac.uk/expressionprofiler • An online microarray data analysis platform What can you do with the data? What can you do with the data? .view Expression Profiler as a heatmap Data Viewer Component What can you do with the data? .clusterExpression Profiler the data Hierarchical Clustering Component What can you do with the data? look at GeneOntology enrichment of a selected cluster Expression... Mus musculus 22% Arabidopsis thaliana 19% Total ~ 7000 hybridisations Apis mellifera Danio rerio Gerbera hybrid cultivar Hordeum vulgare Medicago truncatula Pan troglodytes Platichthys flesus Pseudomonas aeruginosa External Application MAGEML Submission support Curation Database Architecture MAGEML XML AE Data Warehouse MIAMExpress Database ArrayExpress Repository Data upload Data download Visualisation . Data Array 2 Data 2 Hyb protocol Sample protocol Scanning protocol Image analysis protocol Transformation protocol Extraction protocol Hybridisations Labelling protocol Experiment SubmissionArray Submission Protocol. extract Sample hybridisation labelled nucleic acid Microarray RNA extract Sample Experiment Gene expression data matrix normalization integration Protocol Protocol Protocol Protocol Protocol Protocol genes Samples Genes Gene. Transcriptomics Patrick Kemmeren European Bioinformatics Institute Genomics Lab, UMC Utrecht mRNA cDNA hybridise to microarray What are microarrays ? Transcriptomics? hybridisation labelled