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2009,10(1):1112117JournalofPlantGeneticResources枣不同器官和组织RNA提取方法的研究1,2,3,3(1,071000;2,071002;3,071001):,RNACTAB,PVPCTABRNACTAB:2%CTAB,4mol/L,100mmol/LTris2HCl(pH810),20mmol/LEDTA(pH810),2%PVP,4%B2,250Lg/mlTrizolCTABSDS-55RNA,RNATrizol,CTAB,CTABSDS-;CTABRNA;RNA;RNA,,:;;;RNAIsolationofTotalRNAforDifferentOrgansandTissuesofZiziphusjujubaMill1ZHAOJin1,LIUZhong2cheng1,DAILi2,LIUMeng2jun2(1CollegeofLifeSciences,AgriculturalUniversityofHebei,Baoding071000;2CollegeofPharmacy,HebeiUniversity,Baoding071002;3ResearchCenterofChineseJujube,AgriculturalUniversityofHebei,Baoding071001)Abstract:TheleavesofZiziphusjujubaMil.lcv.PozaowasusedasmaterialtoestablishtheimprovedCTABmethodfortotalRNAisolation.TheimprovedaspectsincludedthatremovingphenolbyaddingPVPandB2mercap2toethano,limprovingRNAyieldbyaddingCTABandglyeogen.TheoptimalsystemforRNAisolationare:2%CTAB,4mol/Lguanidineisothiocyanate,210mol/LNaC,l100mmol/LTris2HCl(pH810),20mmol/LEDTA(pH810),2%PVP,4%B2mercaptoethanoland250Lg/mlglyeogen.ThemethodsofisolatingtotalRNAfromplanttissues,includingimprovedCTAB,SDS2pheno,lhotborate,guanidineisothiocyanateandtrizolkitmethod,werecomparedandimproved.Accordingtoquality,yieldandisolatedtime,theresultsshowedthattrizolkitwastheoptimalmethodtoisolateRNAfromtissue2cultureplantlets;improvedCTABwastheoptimalmethodtoisolateRNAfromfieldleavesandfruits;improvedCTABandSDS2phenolweresuitabletoisolateRNAfrombarksandrootskins.ImprovedCTABisacommonmethodtoisolateRNAfromallthetissuesofChinesejujube.RNAwouldac2quiredbydifferenttissuesacrrodingdifferentseasons.ThehighestyieldofRNAwasfieldleaves,andthenwaspla2ntlets,barks,fruitsandrootskins.Keywords:Chinesejujube;Organs;Tissues;TotalRNAisolation:2007212221:2008204230:(C2005000267);(2008BAD92B03):,,E2mai:lzhaojind@yahoo.com:,,Te:l031227521251;E2mai:lkjliu@hebau.edu.cn(ZiziphusjujubaMill1)(Rhamnace2ae)(ZiziphusMill1),,RNA10,cDNART2PCRNorthernRNA[1]RNA,[2],RNase,RNA,RNA,,,RNA,RNA[325],RNARNA,1111RNA112RNA11211RNaseRNARNase,:180e5h;DEPC,011%DEPC015%SDS,ddH2O,RNA11212RNATrizolTrizol,RNACTAB[6],,,CTABB2PVP,1Chomczynski[7]RNAGomez[8],RNASDS2[9]RNA1RNATable1TreatmentsoftotalRNAisolationinChineseju2jubebyimprovedCTABmethodNo1CTAB(%)ConcentrationofCTABB2(%)ConcentrationofB2mercaptoethanolPVP(%)ConcentrationofPVP11222125312104142514561410722282259221010242112451224101332214325153210163421734518341011213RNABACKMANDU800OD260OD230OD280,OD260/OD280OD260/OD230RNA110%RNA=OD260@N()@40(Lg/ml)@V()/(g)2211RNACTAB[2],CTAB,CTAB,RNA,,2323,CTAB1%,RNA21153Lg/g,2%3%14175Lg/g13186Lg/g,OD260/OD230210,OD260/OD28011708,1121:RNA2RNATable2ThepurityandyieldofRNAisolatedfromleavesbydifferenttreatmentsofimprovedCTABmethodNO1UVabsorbanceOD260/OD230OD260/OD280(Lg/g)Yield10169561111442516820176561168891918431190431185041613541164791179822811651152641189642313662103001190351518472151351196241618982160681183421512493133501186041217710218629119810211011131101321130015142124159561190651213513211653119267201111431698111803516113151195161195169192162192082103141417517314840119761111791831953411888711192ODRNATheabsorbancesaretheaveragesofmanytimesisolationindividually,CTAB,2%CTAB;2%4%B2RNA,4%RNA,B24%(1);PVP,PVP,RNA,RNA,PVP,RNA(2),RNA,PVP,2%PVPRNA,CTAB,RNA,4RNA,250Lg/ml750Lg/ml,RNA,,RNA,250Lg/ml3RNATable3TheaveragepurityandyieldofRNAisolatedfromleavesbydifferenttreatmentsofimprovedCTABmethodTreatment(Lg/g)AverageyieldOD260/OD230OD260/OD230averagevalueOD260/OD280OD260/OD280averagevalue1%CTAB211537a114283c117086b2%CTAB141755b311692a119458a3%CTAB131861b310289b119297a2%PVP191280a212130c117822b5%PVP171669a215304b118882a10%PVP141564b218830a119136a2%181124a211293b117903b4%161005a219549a119324a5%,Thesmalllettersfollowingthedatewithineachcolumnrespesentthesignificanceat5%leve,lthesameasbelow113104RNATable4EffectofaddingglyeogeninCTABisolationliquidonRNAisolation(Lg/ml)Concentrationofglyeogen(Lg/g)YieldUVabsorbanceOD260/OD280OD260/OD230015164b119674214367b25021101a119810218629a50020159a119762218064a75021137a119867310581a,,CTAB:2%CTAB,4mol/L,100mmol/LTris2HCl(pH810),20mmol/LEDTA(pH810),2%PVP,4%B2,250Lg/mlCTAB212RNA21211RNA,,RNA5,5RNA,OD260/OD2801179~1196,OD260/OD230210,RNA,,RNA,20h,,,RNA;CTABSDS2RNA,;Trizol,1h,55Lg/g,;RNA,,,,RNA55RNATable5ComparitionbetweendifferentRNAisolationmethodsindifferentorgansandtissuesofChinesejujubeOrganIsolationmethodUVabsorbanceOD260/OD280OD260/OD230(Lg/g#FW)Yield(h)Isolationtime1179b1197d42b4Trizol1182b2102cd55a1CTAB1196a3156a21d3SDS21183b2178b26d3151195a4100a32c201168c0146c24c4Trizol---1CTAB1190a2192a35ab3SDS21183b2143b42a3151188ab3105a31bc201184c2103e32b4Trizol1187c2132d39a1CTAB1198ab3194a18d3SDS21193bc3175b23cd3152105a3125c26bc20---4Trizol---1CTAB1190bc3149b13b3SDS21184c3160b22a3151199ab4137a17b20---4Trizol---1CTAB1185ab2192a17b3SDS21182b0143c24a3151190a1108b22ab20ODRNA;/-0RNATheabsorbancesaretheaveragesofmanytimesisolationindividually;/-0:UnsuccessfulextractionRNA32A,28S18S,RNASDS2518S,3518S,RNA,SDS2RNADNA,1141:RNADNADNase,,RNA,TrizolRNA21212RNA,RNase,RNA,TrizolRNA,,,,RNA,RNARNA,RNA;RNA,OD,,,,RNA;CTABSDS2RNA,,3RNA,SDS2,CTABRNA