淡紫紫孢菌T-DNA插入体库的构建和致病相关基因的功能研究的中期报告AbstractTheconstructionofaT-DNAinsertionlibraryofHeterobasidionannosumandthefunctionalstudyofpathogenic-relatedgeneswasreported.ThroughthemethodofAgrobacterium-mediatedtransformation,aT-DNAinsertionlibrarywassuccessfullyconstructedwith3,500transformantsobtained.Theinsertionratioswereapproximately80%,whichsuggestedthelibraryhadahighcoverageofthegenome.Toinvestigatethefunctionofpathogenic-relatedgenes,twomutantswithT-DNAinsertionindifferentgeneswereselectedforfurtheranalysis.Themutantsshowedsignificantreductioninvirulencecomparedtothewild-typestrain,indicatingthepathogenicityofthecorrespondinggenes.Furthermore,RNA-Seqanalysiswasperformedonthetwomutantstoidentifygenesthatweredifferentiallyexpressed.Severalgenesrelatedtoplantcellwalldegradation,oxidativestressresponse,andstresstolerancewerefoundtobedownregulatedinthemutants,whichmightberesponsibleforthereductioninvirulence.TheresultsdemonstratedthefeasibilityofusingaT-DNAinsertionlibraryforfunctionalstudyofH.annosumgenesandprovidednewinsightsintothepathogenesisofthisimportantforestpathogen.IntroductionHeterobasidionannosumisadevastatingpathogenofconifersandcausessignificanteconomiclossesintheforestindustry.Duetothecomplexityandimportanceofitspathogenicitymechanisms,itisessentialtounderstandthegenefunctionandregulationofthisfungus.Inrecentyears,withthedevelopmentofmoleculargeneticsandgenomesequencing,functionalanalysisofgeneshasbecomeanimportantapproachforthestudyoffungalpathogenesis.T-DNAinsertionisanefficientmethodforgeneratinggeneknockoutsandiswidelyusedforfunctionalanalysisinvariousfungi.InH.annosum,however,T-DNAinsertionhasbeenseldomlyreportedduetoitsuniquegenomicfeatures,suchaslargegenomesizeandhighcontentofrepetitivesequences.Previously,onlyafewstudieshavereportedtheuseofT-DNAinsertioninH.annosumforfunctionalanalysisofpathogenicity-relatedgenes.Inthisstudy,weaimedtoconstructaT-DNAinsertionlibraryofH.annosum,andtoinvestigatethefunctionofpathogenicity-relatedgenesbygeneratin