简介：Precisebaseeditingishighlydesiredinplantfunctionalgenomicresearchandcropmolecularbreeding.Inthisstudy,weconstructedarice-codonoptimizedadeninebaseeditor(ABE)-nCas9toolthatinducedtargetedA·TtoG·Cpointmutationofakeysinglenucleotidepolymorphismsiteinanimportantagriculturalgene.Combinedwiththemodifiedsingle-guideRNAvariant,ourplantABEtoolcanefficientlyachieveadeninebaseeditinginthericegenome.

简介：Homeoboxtranscriptionfactorsparticipateinthegrowthanddevelopmentofplantsbyregulatingcelldifferentiation,morphogenesisandenvironmentalsignalresponse.Torevealthefunctionsofthesetranscriptionfactorsinrice,weconstructedtheRNAivectorsofOsHox9,amemberofhomeoboxfamily,andanalyzedthefunctionofOsHox9usingreversegenetics.TheplantheightandtilleringnumberofRNAitransgenicplantsdecreasedcomparedwiththoseofwild-typeplants.Reversetranscription-polymerasechainreactionanalysisshowedthatOsHox9expressionreducedinthetransgenicplantswithphenotypicvariance,whereasthatinthetransgenicplantswithoutphenotypicvariancewassimilartothatinthewild-typeplants.ThisresultsuggeststhatthephenotypesofthetransgenicplantswerecausedbyRNAieffects.Thetissue-specificityofOsHox9expressionindicatedthatitwasexpressedindifferentorgans,withhighexpressioninstemapicalmeristemandyoungpanicles.SubcellularlocationofOsHox9demonstratedthatitwaslocalizedonthecellmembrane.

简介：EliminationoftheCRISPR/Cas9constructsineditedplantsisaprerequisiteforassessinggeneticstability,conductingphenotypiccharacterization,andapplyingforcommercializationoftheplants.However,removaloftheCRISPR/Cas9transgenesbygeneticsegregationandbybackcrossislaboriousandtimeconsuming.WepreviouslyreportedthedevelopmentofthetransgenekillerCRISPR(TKC)technologythatusesapairofsuicidegenestotriggerself-eliminationofthetransgeneswithoutcompromisinggeneeditingefficiency.TheTKCtechnologyenablesisolationoftransgene-freeCRISPR-editedplantswithinasinglegeneration,greatlyacceleratingcropimprovements.Here,wepresentedtwonewTKCvectorsthatshowgreatefficiencyinbotheditingthetargetgeneandinundergoingself-eliminationofthetransgenes.ThenewvectorsreplacedtheCaMV35SpromoterusedinourpreviousTKCvectorwithtworicepromoterstodriveoneofthesuicidegenes,providingadvantagesoverourpreviousTKCvectorundercertainconditions.Thevectorsreportedhereofferedmoreoptionsandflexibilitytoconductgeneeditingexperimentsinrice.

简介：RiceisanimportantfoodcropinChina,andthedevelopmentofhybridriceisacrucialwaytoincreasegrainyield.Thecreationofdual-purposenuclear-sterilelinesfortwo-linehybridbreedinghasbecomevitalforcommercialricebreeding.WeconstructedthepC1300-2x35S::Cas9-sgRNAPTGMS2-1expressionvectorforeditingthemalefertilitygenePTGMS2-1intwowidelycompatiblericevarieties,93-11andHuazhan,byusingtheCRISPR/Cas9system.Weobtainedthemarker-freephotoperiod-/thermo-sensitivegenicmale-sterile(P/TGMS)linesinT1generation.Accordingtotheexperimentsinphytotronwithfourtemperatureandphotoperiodtreatments,wefoundthetemperatureisthemainfactorforrestoringthepollenfertilityofptgms2-1mutantsin93-11andHuazhan,andthephotoperiodalsohassomeeffectsonpollenfertilityintwodifferentricebackgrounds.Theapplicationofcultivatingnewmale-sterilelinesbygenomeeditingsystemwillsignificantlyacceleratethericebreedingprocess.