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1、RNA-sequencingfromsinglenucleia,1a,2b,2aRashelV.Grindberg,JoyclynL.Yee-Greenbaum,MichaelJ.McConnell,MarkNovotny,a,3cdeaaAndyL.O’Shaughnessy,GeorginaM.Lambert,MarcosJ.Araúzo-Bravo,JunLee,MaxFishman,GillianE.Robbins,fgach,4a,4XiaoyingLin,PratapVenepally,JonathanH.Badger,DavidW.Galbrai
2、th,FredH.Gage,andRogerS.LaskenabJ.CraigVenterInstitute,SanDiego,CA92121;DepartmentofBiochemistryandMolecularGenetics,UniversityofVirginiaSchoolofMedicine,cdCharlottesville,VA22908;SchoolofPlantSciencesandBIO5Institute,UniversityofArizona,Tucson,AZ85721-0036;DepartmentofCellandDevelo
3、pmentalefBiology,MaxPlanckInstituteforMolecularBiomedicine,48149Münster,Germany;LeGeneBiosciences,SanDiego,CA92126;AppliedBiosystems,LifeghTechnologies,FosterCity,CA94404;J.CraigVenterInstitute,Rockville,MD20850;andSalkInstituteforBiologicalStudies,LaJolla,CA92037-1002ContributedbyF
4、redH.Gage,October23,2013(sentforreviewAugust12,2013)Ithasrecentlybeenestablishedthatsynthesisofdouble-strandednucleotideresolution(19).Here,wereportwholetranscriptomecDNAcanbedonefromasinglecellforuseinDNAsequencing.sequencingfromasinglenucleus.TranscriptlevelsinnuclearandGlobalgene
5、expressioncanbequantifiedfromthenumberoftotalcellularRNAfromamouseneuralprogenitorcell(NPC)readsmappingtoeachgene,andmutationsandmRNAsplicinglineweregenerallysimilar,withonlyasmallminoritydifferinginvariantsdeterminedfromthesequencereads.Herewedemon-abundancebetweenthenucleusandthecy
6、toplasm.Currently,stratethatthismethodoftranscriptomicanalysiscanbedoneusingsingle-celltranscriptomicsfromtissuesrequiresproteolyticdis-theextremelylowlevelsofmRNAinasinglenucleus,isolatedfromsociationofcellsatelevatedtemperatureswhichcouldpotentiallyamouseneuralprogenitorcelllinean
7、dfromdissectedhippocam-perturbtranscriptionalactivity.Alternatively,lasercapturemi-paltissue.Thismethodischaracterizedbyexcellentcoverageandcrodissectionmightbeusedtocapturesinglecells(20),however,technicalreproducibility.Onaverage,morethan16,000ofthe24,057mechanicaldamagetothecella
8、nditsdendriticandax
