DNA Methylation Protocols PDF
Preview DNA Methylation Protocols
Methods in Molecular Biology 1708 Jörg Tost Editor DNA Methylation Protocols Third Edition M M B ETHODS IN OLECULAR IO LO GY SeriesEditor JohnM.Walker School of Lifeand MedicalSciences University ofHertfordshire Hatfield, Hertfordshire,AL109AB,UK Forfurther volumes: http://www.springer.com/series/7651 DNA Methylation Protocols Third Edition Edited by Jörg Tost Laboratory for Epigenetics and Environment Centre National de Recherche en Génomique Humaine, CEA—Institut de Biologie Francois Jacob Evry, France Editor Jo¨rgTost LaboratoryforEpigeneticsandEnvironment CentreNationaldeRechercheenGe´nomiqueHumaine CEA—InstitutdeBiologieFrancoisJacob Evry,France ISSN1064-3745 ISSN1940-6029 (electronic) MethodsinMolecularBiology ISBN978-1-4939-7479-5 ISBN978-1-4939-7481-8 (eBook) https://doi.org/10.1007/978-1-4939-7481-8 LibraryofCongressControlNumber:2017960297 ©SpringerScience+BusinessMedia,LLC2018 Thisworkissubjecttocopyright.AllrightsarereservedbythePublisher,whetherthewholeorpartofthematerialis concerned,specificallytherightsoftranslation,reprinting,reuseofillustrations,recitation,broadcasting,reproduction onmicrofilmsorinanyotherphysicalway,andtransmissionorinformationstorageandretrieval,electronicadaptation, computersoftware,orbysimilarordissimilarmethodologynowknownorhereafterdeveloped. Theuseofgeneraldescriptivenames,registerednames,trademarks,servicemarks,etc.inthispublicationdoesnotimply, evenintheabsenceofaspecificstatement,thatsuchnamesareexemptfromtherelevantprotectivelawsandregulations andthereforefreeforgeneraluse. Thepublisher,theauthorsandtheeditorsaresafetoassumethattheadviceandinformationinthisbookarebelievedto betrueandaccurateatthedateofpublication.Neitherthepublishernortheauthorsortheeditorsgiveawarranty, expressorimplied,withrespecttothematerialcontainedhereinorforanyerrorsoromissionsthatmayhavebeenmade. Thepublisherremainsneutralwithregardtojurisdictionalclaimsinpublishedmapsandinstitutionalaffiliations. Covercaption:Mergedimmunostainingofamousebrainfor5-methylcytosine,single-strandedDNAandtotalDNA– ImagetakenfromChapter4ofthisvolume:“AntibodyBasedDetectionofGlobalNuclearDNAMethylationinCells, TissueSectionsandMammalianEmbryos”bySariPenningsandNathalieBeaujeanetal. Printedonacid-freepaper ThisHumanaPressimprintispublishedbySpringerNature TheregisteredcompanyisSpringerScience+BusinessMedia,LLC Theregisteredcompanyaddressis:233SpringStreet,NewYork,NY10013,U.S.A. Preface It is my great pleasure to introduce the third edition of DNA Methylation: Methods and Protocolstothescientificandmedicalcommunity. DNA methylation at cytosines forms one of the multiple layers of epigenetic mechan- isms controlling and modulating gene expression through chromatin structure. It closely interactswithhistonemodificationsandchromatinremodelingcomplexestoformthelocal genomic and higher-order chromatin landscape. DNA methylation is essential for proper mammalian development, crucial for imprinting, and plays a role in maintaining genomic stability.DNAmethylationpatternsaresusceptibletochangeinresponsetoenvironmental stimuli whereby the epigenome seems to be most vulnerable during early life. Changes of DNA methylation patterns have been widely studied in several diseases, especially cancer, where interest has focused on biomarkers for early detection of cancer development, accuratediagnosis, and response to treatment, but havealso been shown to occur in many othercomplexdiseases.Recentadvancesinepigenomeengineeringtechnologiesallownow for the large-scale assessment of the functional relevance of DNA methylation. As a stable nucleicacid-basedmodificationthatistechnicallyeasytohandleandwhichcanbeanalyzed with great reproducibility and accuracy by different laboratories, DNA methylation is a promisingbiomarker for manyapplications. Althoughbynomeanscomplete,thiseditionofDNAmethylation:MethodsandProto- colsgivesacomprehensiveoverviewofavailabletechnologiestogether withadetailedstep- by-step protocol for all experimental procedures required to successfully perform DNA methylationanalysis.Asthedegreeofdifficultyassociatedwithperformingthesemolecular assays is nowadays generally outweighed by the challenges associated with their analysis, manychaptershavethereforealsoincludeddetailedprotocolsfor theanalysisofdatadown tothedetailsofthecommandlinestobeused. ThisisthethirdeditionoftheDNAmethylationprotocols,andthefieldhas—again— dramaticallychangedsincethesecondedition,whichIeditedsixyearsago.DNAmethyla- tiontechnologiesandourknowledgeofDNAmethylationpatternshavebeenadvancingat abreathtakingpaceoverthepastfewyears,andmanyimportantdiscoverieshavebeenmade duetotechnologicaladvances.Mostofthetechniquesdescribedinthesecondeditionhave beenfurtheroptimizedand/orreplacedbynoveleasier,refined,and/ormorequantitative andresolutivetechnologies,manyofwhichcannowbe(andhavealreadybeen)performed on large cohorts. I have therefore again entirely remodeled and expanded the contents of thisbook,whichinthisthirdeditionconsistsnowof35chapters.Onlythreechaptershave beenretainedfromthelastedition,andthesehavebeencompletelyrewrittenbytheauthors to accommodate the changes and improvements made in the last years (Pyrosequencing, MethyLight,andtheHELPassays).Theselectionofdifferenttechnologiespresentedinthis volume enables the analysis of the global DNA methylation content as well as precise quantification of DNA methylation levels on single CpG positions. Methods for the high- resolutionanalysisofCpGpositionswithinatargetregionidentifiedbyoneofthemultiple available genome-wide technologies are presented, and many of the technologies included in a recent international study comparing the reliability and performance of locus-specific DNAmethylationassays(Bocketal.,Nat.Biotech.2016)areincludedinthisvolume.While the second edition contained a large number of microarray-based analysis methods, they v vi Preface have been completely superseded by sequencing-based approaches. With sequencing costs further decreasing, whole genome bisulfite sequencing (WGBS) will soon become afford- able in large cohorts and several protocols with different approaches such as MethylC-seq, PBAT, or tagmentation-based WGBS are presented in this volume. Special focus has been given to protocols that enable to start with low amounts of starting material allowing to analyzethecell-typeordevelopmental-stage-specific DNAmethylationpatterns.Genome- wide sequencing approaches will lead to a large number of potential candidate genes, and therefore this volume now includes a novel section detailing methods analyzing a large number of target regions in parallel and the interested reader can choose from a variety of captureoramplification-basedtechnologies. Forthefirsttime,achapterisdedicatedtotheexperimentaldesignofDNAmethylation studies as well as the statistical considerations on their analysis, while a second chapter focuses onanalysis strategiesfor thehighly popularIllumina BeadArrayswith thedifferent parameters to be considered during data analysis and interpretation. Analysis of cell-free DNA at both the genetic and epigenetic level has attracted recently much interest for the diagnosis and clinical management of cancer and other complex diseases, and this volume provides a protocol for the DNA methylation analysis in body fluids. This collection also contains a protocol for DNA methylation analysis from archived Guthrie cards potentially enabling longitudinal studies and identifying DNA methylation changes prior to disease onset.Withtheincreasinginterestinanalyzingseveralepigeneticmodificationsonthesame samples, this volume contains also two chapters describing the integrated analysis of DNA methylationpatternsinimmunoprecipitatedDNAandthusassociatedwithspecifichistone modificationsortranscriptionfactorprofilesaswellaswithnucleosomeoccupancy.Asinthe last edition, an introductory chapter summarizes briefly the different biological processes DNAmethylationisinvolvedin,thechangesthathavebeendescribedindiseasesaswellas potential clinical applications for which DNA methylation analysis might prove to be important allowing scientists to catch up with the current level of knowledge and learn aboutrecenttrends. ThisvolumeoftheMethodsinMolecularBiologyseriescontainswidelyusedmethods such as Pyrosequencing and methylation-specific PCR as well as protocols for special applications suchas thehairpin bisulfitesequencing whichallows for theassessment ofthe symmetry of DNA methylation in specific regions. Furthermore, research of the last years has shown that cytosine methylation is not the only DNA modification and that especially 5-hydroxymethylation is not only an intermediate in oxidative DNA demethylation, but constitutes a distinct layer in the complex process of epigenetic regulation with its own distributionandregulatoryfunctions.Asbisulfite-basedtechnologiesarenotabletodistin- guishbetweenthesetwomodifications,specialprotocolshavebeendevelopedandseveralof themarepresentedinthelastsectionofthisbook. This book is addressed to postdoctoral investigators and research scientists that are implicatedinthedifferentaspectsofgeneticsandcellularandmolecularbiologyaswellasto clinicians involved in diagnostics or choice of treatment of diseases that have an epigenetic component,whichnowadaysmeansmostbiologicalandclinicalquestions.Thepresentation inthisvolumeisequallysuitedforlaboratoriesthatalreadyhaveagreatdealofexpertiseina certain technology to analyze DNA methylation, but might want to obtain other or complementary data using an orthogonal technique, and for genetics/genomics/biology groups that want to initiate research in this exciting area and want to identify the method best suited to answer their question. Notes and tips from experts of the different methods will enable a rapid implementation of the different protocols in the laboratory and avoid Preface vii time-consuming and cost-intensive mistakes. With the tools and protocols available, our knowledge and understanding of DNA methylation will continue to increase rapidly with new groundbreaking and exciting discoveries to come, and this book will contribute to spreadingofthe“savoir faire”toanalyzeDNAmethylation. I am indebted to all the authors for their hard work and outstanding contributions to thisthirdeditionofDNAMethylationProtocols.Itwasapleasuretoworkwiththemonthis project.Ihopethattheprotocolsdescribedindetailinthisvolumewillhelptoacceleratethe analysisanddescriptionofthe“methylome”ofdifferentspecies,enhanceourunderstanding ofthemolecularprocessesthatdeterminethegenomicDNAmethylationlandscapeaswell as provide robust technologies for the clinical implementation of DNA methylation-based biomarkers. Evry,France J¨orgTost Contents Preface ..................................................................... v Contributors................................................................. xiii PART I INTRODUCTION 1 ASummaryoftheBiologicalProcesses,Disease-AssociatedChanges, andClinicalApplicationsofDNAMethylation ............................. 3 GitteBrinchAndersenandJ¨orgTost 2 ConsiderationsforDesignandAnalysisofDNAMethylationStudies ......... 31 KarinB.MichelsandAlexandraM.Binder PART II GLOBAL DNA METHYLATION LEVELS 3 QuantificationofGlobalDNAMethylationLevelsbyMass Spectrometry........................................................... 49 AgustinF.Fernandez,LuisValledor,FernandoVallejo,MariaJesu´sCan˜al, andMarioF.Fraga 4 Antibody-BasedDetectionofGlobalNuclearDNAMethylationinCells, TissueSections,andMammalianEmbryos................................. 59 NathalieBeaujean,JulietteSalvaing,NurAnniesAbdHadi, andSariPennings PART III GENOME-WIDE DNA METHYLATION ANALYSIS ® 5 Whole-GenomeBisulfiteSequencingUsingtheOvation Ultralow Methyl-SeqProtocol .................................................... 83 ChristianDaviaud,VictorRenault,FlorenceMauger,Jean-Franc¸ois Deleuze,andJ¨orgTost 6 Tagmentation-BasedLibraryPreparationforLowDNAInput WholeGenomeBisulfiteSequencing...................................... 105 DieterWeichenhan,QiWang,AndrewAdey,StephanWolf,JayShendure, RolandEils,andChristophPlass 7 Post-BisulfiteAdaptorTaggingforPCR-FreeWhole-GenomeBisulfite Sequencing............................................................. 123 FumihitoMiuraandTakashiIto 8 MultiplexedReducedRepresentationBisulfiteSequencingwithMagnetic BeadFragmentSizeSelection ............................................ 137 WilliamP.AccomandoJr.andKarinB.Michels 9 LowInputWhole-GenomeBisulfiteSequencingUsingaPost-Bisulfite AdapterTaggingApproach............................................... 161 JulianR.PeatandSe´bastienA.Smallwood ix x Contents 10 Methyl-CpG-BindingDomainSequencing:MBD-seq....................... 171 KarolinaA.Aberg,RobinF.Chan,LinyingXie,AndreyA.Shabalin, andEdwinJ.C.G.vandenOord 11 TheHELP-BasedDNAMethylationAssays................................ 191 JohnM.Greally 12 ComprehensiveWholeDNAMethylomeAnalysisbyIntegrating MeDIP-seqandMRE-seq ............................................... 209 XiaoyunXing,BoZhang,DaofengLi,andTingWang 13 DigitalRestrictionEnzymeAnalysisofMethylation(DREAM)............... 247 JaroslavJelinek,JustinT.Lee,MatteoCesaroni,JozefMadzo, ShoudanLiang,YueLu,andJean-PierreJ.Issa 14 NucleosomeOccupancyandMethylomeSequencing(NOMe-seq) ........... 267 FidesD.Lay,TheresaK.Kelly,andPeterA.Jones 15 BisulphiteSequencingofChromatinImmunoprecipitatedDNA (BisChIP-seq).......................................................... 285 ClareStirzaker,JennyZ.Song,AaronL.Statham,andSusanJ.Clark 16 AGuidetoIlluminaBeadChipDataAnalysis............................... 303 MichaelC.WuandPei-FenKuan PART IV ANALYSIS OF HIGHLY MULTIPLEXED TARGET REGIONS 17 MicrodropletPCRforHighlyMultiplexedTargetedBisulfite Sequencing ............................................................ 333 H.KiyomiKomori,SarahA.LaMere, TraverHart,StevenR.Head, AliTorkamani,andDanielR.Salomon 18 MultiplexedDNAMethylationAnalysisofTargetRegionsUsing Microfluidics(Fluidigm)................................................. 349 MartynaAdamowicz,KlioMaratou,andTimothyJ.Aitman 19 Large-ScaleTargetedDNAMethylationAnalysisUsingBisulfite PadlockProbes......................................................... 365 DinhDiep,NonglukPlongthongkum,andKunZhang 20 TargetedBisulfiteSequencingUsingtheSeqCapEpiEnrichment System................................................................. 383 JenniferWendt,HeidiRosenbaum,ToddA.Richmond,Jeffrey A.Jeddeloh, andDanielL.Burgess 21 MultiplexedandSensitiveDNAMethylationTestingUsing Methylation-SensitiveRestrictionEnzymes“MSRE-qPCR” ................. 407 GabrielBeikircher,WalterPulverer,ManuelaHofner,ChristaNoehammer, andAndreasWeinhaeusel PART V LOCUS-SPECIFIC DNA METHYLATION ANALYSIS 22 QuantitativeDNAMethylationAnalysisatSingle-NucleotideResolution ® byPyrosequencing ..................................................... 427 FlorenceBusato,EmelyneDejeux,HafidaElabdalaoui,IvoGlynneGut, andJ¨orgTost Contents xi 23 Methylation-SpecificPCR................................................ 447 Joa˜oRamalho-Carvalho,RuiHenrique,andCarmenJer(cid:1)onimo 24 QuantitationofDNAMethylationbyQuantitativeMultiplex Methylation-SpecificPCR(QM-MSP)Assay............................... 473 MaryJoFacklerandSaraswatiSukumar 25 MethyLightandDigitalMethyLight ...................................... 497 MihaelaCampan,DanielJ.Weisenberger,BinhTrinh,andPeterW.Laird 26 QuantitativeRegion-SpecificDNAMethylationAnalysisbythe EpiTYPER™Technology................................................ 515 SonjaKunze 27 Methylation-SpecificMultiplexLigation-DependentProbeAmplification (MS-MLPA) ........................................................... 537 CathyB.Moelans,LilitAtanesyan,SuviP.Savola,andPaulJ.vanDiest 28 Methylation-SensitiveHighResolutionMelting(MS-HRM)................. 551 DiannaHussmannandLiseLotteHansen 29 HairpinBisulfiteSequencing:SynchronousMethylationAnalysis onComplementaryDNAStrandsofIndividualChromosomes............... 573 PascalGiehrandJ¨ornWalter 30 Helper-DependentChainReaction(HDCR)forSelectiveAmplification ofMethylatedDNASequences........................................... 587 SusanM.Mitchell,KeithN.Rand,Zheng-ZhouXu,ThuHo, GlennS.Brown,JasonP.Ross,andPeterL.Molloy PART VI DNA METHYLATION ANALYSIS OF SPECIFIC BIOLOGICAL SAMPLES 31 DNAMethylationAnalysisfromBloodSpots:IncreasingYield andQualityforGenome-WideandLocus-SpecificMethylationAnalysis....... 605 AkramGhantous,HectorHernandez-Vargas,andZdenkoHerceg 32 DNAMethylationAnalysisofFree-CirculatingDNAinBodyFluids.......... 621 MariaJung,GlenKristiansen,andDimoDietrich PART VII HYDROXYMETHYLATION 33 Tet-AssistedBisulfiteSequencing(TAB-seq) ............................... 645 MiaoYu,DaliHan,GaryC.Hon,andChuanHe 34 MultiplexingforOxidativeBisulfiteSequencing(oxBS-seq).................. 665 KristinaKirschner,FelixKrueger,AnthonyR.Green,andTamirChandra 35 Affinity-BasedEnrichmentTechniquesfor theGenome-Wide Analysisof5-Hydroxymethylcytosine...................................... 679 JohnP.ThomsonandRichardR.Meehan Index ...................................................................... 697
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