抗黄瓜花叶病毒RNAi载体的构建及烟草的转化(5)

2021-09-24 20:08

7.5NS04CMV2a91.972.091.693.691.694.887.595.271.894.791.991.491.492.193.493.495.08

Thedown?leftisnucleotideidentities,theupper?rightisaminoacididentities.1-19arecorrespondingwiththesequenceNo.intheleftoffigurefromuptobottom

M:DL2000;1:CR2?BKsequence.Fig.3 PCRamplificationofCR2?B

M:DL4500;1:piRDCR2.SwithClaI.

Fig.5 Therestrictionenzymedigestionofrecombinantplas?

midpiRDCR2.

M:DL2000;1:CR2?BKsequence.

Fig.4 PCRamplificationofCR2?SB

ObtainingoftransgenictobaccoplantsTransgenictobaccoplantsDNAwasextracted,andthentheamplificationwascarriedoutbyprimersCR2Kp/CR2SB

,the1.2%ofagarosegelandCR2Bg/CR2SBrespectively

,itcouldbeconcludedelectrophoresiswasusedforanalysis

thatthePCR_CR2_BKfragmentwith693bpandthePCR_CR2_SBfragmentwith741bpwereamplified(Fig.7,8)

M:DL4500;1:pBi35SCR2withBamHⅠ.

Fig.6 Therestrictionenzymedigestionofrecombinantplas?

midpBi35SCR2

indicatingthattheobtainedtobaccoplantshadbeensuccess?

fullytransformedintothepBi35SCR2expressionvectorAgrobacterium?mediatedmethod.

72AgriculturalScience&TechnologyVol.11,No.5,2010

[10]

.Theconstructedexpres?theefficiencyofgenesilencing

icotianato?sionvectorpBi35SCR2wastransformedintotheN

baccowithAgrobacterium?mediatedmethod,theobtainedtransgenictobaccoplantscouldbeobservedontheantiviralabilityoftransgenictobaccoplantsbychallengetest,ifthe

,itcouldbefurtherappliedtotomatopro?effectwasobvious

cessing,whichhadlaidthefoundationfortheanti?CMVpro?cessingtomatoplants.

M:DL2000;1:pBi35SCR2vectorcontrol;2-5:pBi35SCR2vec?

tortransgenictobaccolines;6:PBI121transgenictobacco.Fig.7 PCRidentificationofCR2_BKfragmentintransgenicto?

baccoplant

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(heritanceofcucumbermosaicviruslmnopqprstuvw

[J].TianjinAgriculturalSciences(,2008,14z{|},-)xy)(2):55-58.[2]EAMENSA,WANGMB,SMITHNA,etal.RNAsilencingin

:yesterday,today,andtomorrow[J].PlantPhysiol,2008,plants147(2):456-468.[3]MATZKEM,MATZKEAJ,KOOTERJM.RNA:guidinggenesilen?

[J].Science,2001,293(5532):1080-1083.cing

[4]ZAMOREPD,TUSCHLT,SHARPPA,etal.RNAi:double?stran?

dedRNAdirectstheATP?dependentcleavageofmRNAat21to23

J].Cell,2000,101:25-33.nucleotideintervals[

[5]SIJENT,KOOTERJM.Post?transcriptionalgenesilencing:RNAs

ontheattackoronthedefense?[J].Bioessays,2000,22(6):520-531.[6]SHARPPA.RNAinterference[J].Genes&Development,2001,

15(5):485-490.[7]GUOXQ(,LUSE(,ZHUCX(,etal.~ ) ) )

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[J].ActaPhytopathologicaSinica( YpqZ ) Fp

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