Cap-Analysis Gene Expression (CAGE) The Science of Decoding Genes Transcription

A guide for users of new technologies, this volume includes accurately proven protocols, allowing readers to prepare their samples for experiments. Additionally, it provides a guide for the bioinformatics tools that are available for the analysis of the obtained tags, including the design of the software, the sources and web information where they can be downloaded. Finally, the book provides examples of the application of these technologies to identify promoters, annotate genomes, identify new RNAs and reconstruct models of transcriptional control. Although examples mainly regard mammalians, the discussion expands to other groups of eukaryotes, where these approaches are complementing genome sequencing.

This book is a guide for users of new technologies, as it includes accurately proven protocols, allowing readers to prepare their samples for experiments. Additionally, it is a guide for the bioinformatics tools that are available for the analysis of the obtained tags, including the design of the software, the sources and the Web. Finally, the book provides examples of the application of these technologies to identify promoters, annotate genomes, identify new RNAs and reconstruct models of transcriptional control. Although examples mainly concern mammalians, the discussion expands to other groups of eukaryotes, where these approaches are complementing genome sequencing.

Preface vCap Analysis Gene Expression (CAGE)Tagging Transcription Starting Sites with CAGEThe Output of the Genome is ComplexMapping 5’ Ends: From ESTs to Tagging TechnologiesLinking Core Promoters to Genomic ElementscDNA Ends or the Whole Sequence?Identification of Functional Elements in the GenomeTechnology Evolution, Same Lessons?Construction of CAGE LibrariesIntroductionStage 1: Synthesis of First-Strand cDNAStage 2: Oxidation/BiotinylationStage 3: Capture-ReleaseStage 4: Single Strand Linker LigationStage 5: the Second Strand cDNA SynthesisStage 6: Preparing CAGE TagsStage: 7 Amplification of CAGE TagsStage 8: RestrictionStage 9: ConcatenationTranscriptome and Genome Characterization Using Massively Parallel Paired End Tag (PET) Sequencing AnalysisIntroductionThe Development of Pair end diTag (PET) AnalysisGIS-PET for Transcriptome AnalysisChIP-PET for Whole Genome Mapping of Transcription Factor Binding Sites and Epigenetic ModificationsChIA-PET for Whole Genome Identification of Long Range interactionsPerspectiveNew Era of Genome-Wide Gene Expression AnalysisIntroductionTagging Technologies for Genome-Wide AnalysisPrinciples of Next Generation Sequencing TechnologiesGenome Analyzer (Illumina/Solexa)SOLiD System (Applied Biosystems)Advantages of Next Generation Sequencing Technologies over Conventional Sequencing Technology on Tagging TechnologiesFrom Static Analysis to Dynamic AnalysisCAGE Method and Next Generation Sequencing TechnologiesConclusions and OutlookComputational Tools to Analyze CAGE Introduction to PART II Extraction and Quality Control of CAGE TagsOverviewUsing Read Qualities and Read Properties, Pre- and Post-ExtractionProcedures Before Tag ExtractionUsing QC Values After Tag ExtractionOrigin of Sequence ErrorsUsing Sequence Errors to Estimate CAGE QualityA Simple CAGE Tag Extraction MethodSetting CAGE Tags in a Genomic ContextMapping Pipelines for Sequence Tag TechnologiesA Mapping Pipeline for CAGEBenchmarking with a Sample DatasetUsing CAGE Data for Quantitative ExpressionHigh Throughput Expression PlatformsComparing CAGE to Other Measures of Gene ExpressionPlatform NormalizationReplicationGene Models and Complex LociConstruction of CAGE Promoters and Calculation of Gene Expression LevelsComparison of CAGE Expression between Technical ReplicatesComparison of CAGE Expression from Biological ReplicatesComparison of CAGE Expression Between Different Time Points Within a Single Time-CourseComparison of CAGE Expression Profiling to qRT-PCR Expression MeasurementsComparison of CAGE Expression Profiling to MicroarrayMeasurementsPresent/Absent CallsDiscussionDatabases for CAGE Visualization and AnalysisIntroductionTranscription Maps and ActivityPublic DatabasesGenomic View of In-House DataFor Expression AnalysesDiscussionComputational Methods to Identify Transcription Factor Binding Sites Using CAGE InformationIntroductionSchema of the Methodology ProcessInitial Links of TF with the Affected GenesCorrelation of CAGE Tag Counts of Genes and TFsRanking TF!TFBS!TSS/Promoter!GENE Association: The Effective Use of CAGE TagsVerification of ResultsReconstruction of TRNsTranscription Regulatory Networks Analysis Using CAGECAGE Data for Network ReconstructionMethodologyGene Expression Data Complementary to CAGE for Network ReconstructionUsing Physical InteractionsTRNs ReconstructionUsing Pathway InformationValidation of the Reconstructed NetworksGene-Expression Ontologies and Tag-Based Expression ProfilingIntroductionAnnotating Gene Expression Using Ontologies to Integrate Expression InformationLessons Learned from Genomic CAGEIntroductionThe Classic View on Transcription Start Sites and Core PromotersCAGE-Based Views of Transcription Start SitesProbing Biological Mechanisms Using CAGEFuture Challenges in CAGE AnalysisWhat are we Measuring?How Close to "The Truth" areWe?Comparative Genomics and Mammalian Promoter Evolution IntroductionResources for Comparative Genomic AnalysisGenome Wide Trends in Mammalian Promoter EvolutionPromoters Represent an Unusual Genomic EnvironmentIntegration of Population Genetic Data with Comparative GenomicsConcluding RemarksThe Impact of CAGE Data on Understanding Macrophage Transcriptional BiologyIntroductionTranscription start site and promoter characteristics revealed by CAGETranscriptional Complexity: Sense-Antisense Transcription and Non-Coding RNAConstruction of Macrophage Transcriptional NetworksWhat does CAGE Data Offer for Traditional Studies of Promoter Regulation?ConclusionColor IndexIndex

Piero Carninci was born in Italy in 1965 and earned his doctorate in biological science at the University of Trieste in 1989. For six months from April 1989, he worked as a research associate at the International Center for Genetic Engineering and Biotechnology and then served as a health assistant in the Italian army. In 1990, he became a researcher at a biotech company called TALENT, after which he moved to RIKEN as a researcher in 1995. Since 2003, he has been working as a senior scientist at the Genome Science Laboratory at the RIKEN Frontier Research Institute and at the RIKEN Genomic Sciences Center.