Page 23 - 《华农农业大学学报》2020年第3期

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华 中 农 业 大 学 学 报 第 ３９ 卷
Molecular p lant , ２０１７ , １０ ( ７ ): ９１８Ｇ９２９． co p e p tidere p eatp roteincausesaseedlin g Ｇs p ecificalbinop heＧ
[ ２１ ] LIU H , DING Y , ZHOU Y , etal．CRISPRＧP２．０ : anim p roved not yp eanditsutilizationtoenhanceseedp urit y inh y bridrice
CRISPRＧCas９toolfor g enomeeditin g inp lants [ J ] ．Molecular p roduction [ J ] ．Plant p h y siolo gy , ２０１２ , １５９ ( １ ): ２２７Ｇ２３８．
p lant , ２０１７ , １０ ( ３ ): ５３０Ｇ５３２． [ ２８ ] TOMINAGAJ , MIZUTANI H , HORIKAWA D , etal．Rice
[ ２２ ] XIAO H , XU Y , NIC , etal．A ricedualＧlocalizedp entatriＧ CYO１ , anortholo gofArabido p sisthalianacot y ledonchloroＧ
co p e p tidere p eat p roteinisinvolvedinor g anellarRNAeditin g p lastbio g enesisfactorAtCYO１ , isex p ressedinleavesandinＧ
to g etherwithOsMORFs [ J ] ．Journalofex p erimentalbotan y , volvedin p hotos y ntheticp erformance [ J ] ．Journalofp lant
２０１８ , ６９ ( １２ ): ２９２３Ｇ２９３６． p h y siolo gy , ２０１６ , ２０７ : ７８Ｇ８３．
[ ２３ ] HEL , ZHANGS , QIUZ , etal．FRUCTOKINASEＧLIKEPROＧ [ ２９ ] PEREZＧRUIZJM , SPINOLA M C , KIRCHSTEIGERK , etal．
TEIN１interactswithTRXztore g ulatechloro p lastdevelo p Ｇ RiceNTRCisahi g hＧefficienc yredoxs y stemforchloro p last
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( ２ ): ９４Ｇ１１１． ( ９ ): ２３５６Ｇ２３６８．
[ ２４ ] MAX , SUNX , LIC , etal．Ma p Ｇbasedclonin g andcharacterizaＧ [ ３０ ] DONG H , FEIGL , WUCY , etal．AricevirescentＧ y ellowleaf
tionofthenovely ellowＧ g reenleafg eneYS８３inrice ( Or y za mutantrevealsnewinsi g htsintotheroleandassembl y of p lasＧ
sativa )[ J ] ．Plant p h y siolo gyandbiochemistr y , ２０１７ , １１１ : １Ｇ９． tidcaseinol y ticp roteaseinhi g her p lants [ J ] ．Plant p h y siolo gy ,
[ ２５ ] CHEN H , CHENGZ , MA X , etal．Aknockdownmutationof ２０１３ , １６２ ( ４ ): １８６７Ｇ１８８０．
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[ ２６ ] WANGY , ZHANGJ , SHIX , etal．Tem p eratureＧsensitivealbiＧ [ J ] ．Plantscience , ２０１２ , １９５ : １２５Ｇ１３４．
nog eneTCD５ , encodin gamonoox yg enase , affectschloro p last [ ３２ ] HOUX , XIEK , YAOJ , etal．Ahomolo g ofhumanskiＧinteracＧ
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botan y , ２０１６ , ６７ ( １７ ): ５１８７Ｇ５２０２． tolerance [ J ] ．Proceedin g softhe National Academ yofSciＧ
[ ２７ ] SU N , HU M L , WU DX , etal．Disru p tionofaricep entatriＧ ences , USA , ２００９ , １０６ ( １５ ): ６４１０Ｇ６４１５．

,
Asim p lefastandefficientmethodofconstructin g
dualＧtar g etCRISPR Cas９vector
/

,
,
ZHANG Yan g HUANG Weifen gZHOUFei , LIN Yon gj un
Colle g eo fBiolo gyScienceandTechnolo gy NationalKe yLaborator yo fCro pGeneticIm p rovement ,
/
Huazhon gA g riculturalUniversit y Wuhan４３００７０ , China
,

Abstract Asim p le , fastandefficientmethodofconstructin gdualＧtar g etCRISPR / Cas９vectorwas
establishedbasedontheGoldenGateli g ation．Thetwodesi g nedsmalltar g etDNAfra g mentsandone
exo g enousfra g mentwereconstructedto g etherintotheCRISPR / Cas９vectorwithT ４DNAli g ase．２０duＧ
alＧtar g etCRISPR / Cas９vectorsa g ainst１０re p ortedrice g eneswereconstructedwiththismethod , andthe
,
p ositiveratewas１００％ withnomutationinse q uences．Thisvectors y stemp rovidesaneas yra p idandefＧ
ficientmethodforconstructin gdualＧtar g etCRISPR / Cas９vectorforrice．Italsop rovidesareferencefor
theconstructionofdualＧtar g etCRISPR / Cas９vectorsinother p lants．
;
g
g
Ke y words rice ; CRISPR / Cas９ ; eneknockout ; eneeditin g dualＧtar g et ; vectorconstruction
( 责任编辑: 张志钰)