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犆犾狅狀犲犪狀犱犈狓狆狉犲狊狊犻狅狀犃狀犪犾狔狊犻狊狅犳犌犌犘犘犛1犌犲狀犲犳狉狅犿犚犲犺犿犪狀狀犻犪犵犾狌狋犻狀狅狊犪
ZHAOLe1,2,SHIJingjing1,2,MALigang1,2,HEQingxiang1,FENGWeisheng1,2,
ZHENGXiaoke1,2,ZHUYunhao1,2
(1SchoolofPharmacy,HenanUniversityofTraditionalChineseMedicine,Zhengzhou450046,China;2ColaborativeInnova
tionCenterforRespiratoryDiseaseDiagnosisandTreatment&ChineseMedicineDevelopmentofHenanProvince,Zhengzhou
450046,China)
犃犫狊狋狉犪犮狋:犚犲犺犿犪狀狀犻犪犵犾狌狋犻狀狅狊犪wasusedasexperimentalmaterialinthisstudy.Throughanalyzingthe
transcriptomedataof犚.犵犾狌狋犻狀狅狊犪anddesigningspecificprimers,weisolatedthecDNAsequenceofgera
nylgeranylpyrophosphatesynthase(GGPPS)from犚.犵犾狌狋犻狀狅狊犪andnamedas犚犵犌犌犘犘犛1(GenBankac
cessionNo.KU258808).Meanwhile,onthebasisofbioinformaticanalysis,weperformedtheprokaryotic
expression,purificationandtissuespecificexpressionanalysis.Theresultsindicatedthat:(1)犚犵犌犌
犘犘犛1hasanopenreadingframe(ORF)of987bp,whichencodedaproteinof328aminoacidresidues.
(2)BioinformaticanalysisindicatedthatRgGGPPS1proteincontainsthetwoconservedmotifs(DDXXXX
DD)and(DDXXD);RgGGPPS1proteinshowedthehighesthomology,92%identity,withGGPPSpro
teinfrom犛犲狊犪犿狌犿犻狀犱犻犮狌犿.(3)ByutilizingtheconstructionofprokaryoticexpressionvectorpET32a
犚犵犌犌犘犘犛1,wesuccessfulyexpressedtherecombinantRgGGPPS1proteinin犈.犮狅犾犻BL21(DE3)cels.
Furthermore,therecombinantRgGGPPS1proteinwaspurifiedthroughNi2+affinitychromatography.(4)
RealtimePCRanalysisindicatedthat犚犵犌犌犘犘犛1wasexpressedinhightranscriptlevelinroots,lowlev
elsinleavesandstems.Theresultsofthisstudyprovidedthefundamentalinformationabout犚犵犌犌犘犘犛1
geneforfolowupresearchofitsfunctioninvolvediniridoidglycosidebiosynthesispathway.
犓犲狔狑狅狉犱狊:犚犲犺犿犪狀狀犻犪犵犾狌狋犻狀狅狊犪;犚犵犌犌犘犘犛1gene;bioinformaticanalysis;prokaryoticexpression;ex
pressionanalysis
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Fig.1 PCRamplificationof犚犵犌犌犘犘犛1gene
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Themarker“”representsterminationcodon,theleftnumberindicatesnucleotideposition,therightnumberindicates
aminoacidposition,theitaliclowercaselettersrepresentcloningprimersequencesof犚犵犌犌犘犘犛1gene
Fig.2 Nucleotidesequenceandaminoacidsequenceof犚犵犌犌犘犘犛1genecDNA
1985
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drawnfromthebootstraptest
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