JournalofVisualizedExperimentswww.jove.comCopyright©2012CreativeCommonsAttribution-NonCommercialLicenseJuly2012|65|e3938|Page1of7VideoArticleOptimizedAnalysisofDNAMethylationandGeneExpressionfromSmall,Anatomically-definedAreasoftheBrainMarcBettscheider1,ArletaKuczynska1,OsborneAlmeida1,DietmarSpengler11MaxPlanckInstituteofPsychiatryCorrespondenceto:DietmarSpengleratspengler@mpipsykl.mpg.deURL:http://www.jove.com/video/3938DOI:doi:10.3791/3938Keywords:Neuroscience,Issue65,Genetics,Physiology,Epigenetics,DNAmethylation,early-lifestress,maternalseparation,bisulfitesequencingDatePublished:7/12/2012Citation:Bettscheider,M.,Kuczynska,A.,Almeida,O.,Spengler,D.OptimizedAnalysisofDNAMethylationandGeneExpressionfromSmall,Anatomically-definedAreasoftheBrain.J.Vis.Exp.(65),e3938,doi:10.3791/3938(2012).AbstractExposuretodiet,drugsandearlylifeadversityduringsensitivewindowsoflife1,2canleadtolastingchangesingeneexpressionthatcontributetothedisplayofphysiologicalandbehaviouralphenotypes.Suchenvironmentalprogrammingislikelytoincreasethesusceptibilitytometabolic,cardiovascularandmentaldiseases3,4.DNAmethylationandhistonemodificationsareconsideredkeyprocessesinthemediationofthegene-environmentdialogueandappearalsotounderlayenvironmentalprogramming5.Inmammals,DNAmethylationtypicallycomprisesthecovalentadditionofamethylgroupatthe5-positionofcytosinewithinthecontextofCpGdinucleotides.CpGmethylationoccursinahighlytissue-andcell-specificmannermakingitachallengetostudydiscrete,smallregionsofthebrainwherecellularheterogeneityishighandtissuequantitylimited.Moreover,becausegeneexpressionandmethylationarecloselylinkedevents,increasedvaluecanbegainedbycomparingbothparametersinthesamesample.Here,astep-by-stepprotocol(Figure1)fortheinvestigationofepigeneticprogramminginthebrainispresentedusingthe'maternalseparation'paradigmofearlylifeadversityforillustrativepurposes.Theprotocoldescribesthepreparationofmicropunchesfromdifferentially-agedmousebrainsfromwhichDNAandRNAcanbesimultaneouslyisolated,t...
