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1、植物遗传资源学报2009,10(1):1112117JournalofPlantGeneticResources枣不同器官和组织RNA提取方法的研究1233赵锦,刘中成,代丽,刘孟军(1河北农业大学生命科学学院,保定071000;2河北大学药学院,保定071002;3河北农业大学中国枣研究中心,保定071001)摘要:以田间幼嫩叶片为试材,建立枣RNA改良CTAB提取法,改良之处包括利用巯基乙醇和PVP联合去除多酚、CTAB与异硫酸氰胍联合促进RNA释放及加入糖原提高产率等。改良CTAB法提取液组成:2%CTAB
2、,4mol/L异硫酸氰胍,100mmol/LTris2HCl(pH810),20mmol/LEDTA(pH810),2%PVP,4%B2巯基乙醇,250Lg/ml糖原。进而采用Trizol试剂法、改良CTAB法、改进SDS-酚法、热硼酸法和异硫氰酸胍法5种方法分别对枣组培苗、幼叶、幼嫩枝皮、根皮和枣果5种器官和组织进行了总RNA提取。结果表明,组培苗RNA提取以Trizol试剂法为最佳,田间幼叶和成熟果实宜采用改良CTAB法,幼嫩枝皮和根皮宜采用改良CTAB法或改进SDS-酚法;本研究建立的改良CTAB法可作为枣
3、不同器官和组织RNA提取的通用方法;根据季节可从不同器官和组织中获得高质量RNA;枣组织RNA含量相差较大,以幼嫩叶片含量最高,其次为组培苗、枝皮、枣果和根皮。关键词:枣;器官;组织;总RNA提取IsolationofTotalRNAforDifferentOrgansandTissuesofZiziphusjujubaMill11122ZHAOJin,LIUZhong2cheng,DAILi,LIUMeng2jun12(CollegeofLifeSciences,AgriculturalUniversityof
4、Hebei,Baoding071000;CollegeofPharmacy,Hebei3University,Baoding071002;ResearchCenterofChineseJujube,AgriculturalUniversityofHebei,Baoding071001)Abstract:TheleavesofZiziphusjujubaMil.lcv.PozaowasusedasmaterialtoestablishtheimprovedCTABmethodfortotalRNAisolation
5、.TheimprovedaspectsincludedthatremovingphenolbyaddingPVPandB2mercap2toethano,limprovingRNAyieldbyaddingCTABandglyeogen.TheoptimalsystemforRNAisolationare:2%CTAB,4mol/Lguanidineisothiocyanate,210mol/LNaC,l100mmol/LTris2HCl(pH810),20mmol/LEDTA(pH810),2%PVP,4%B2
6、mercaptoethanoland250Lg/mlglyeogen.ThemethodsofisolatingtotalRNAfromplanttissues,includingimprovedCTAB,SDS2pheno,lhotborate,guanidineisothiocyanateandtrizolkitmethod,werecomparedandimproved.Accordingtoquality,yieldandisolatedtime,theresultsshowedthattrizolkit
7、wastheoptimalmethodtoisolateRNAfromtissue2cultureplantlets;improvedCTABwastheoptimalmethodtoisolateRNAfromfieldleavesandfruits;improvedCTABandSDS2phenolweresuitabletoisolateRNAfrombarksandrootskins.ImprovedCTABisacommonmethodtoisolateRNAfromallthetissuesofChi
8、nesejujube.RNAwouldac2quiredbydifferenttissuesacrrodingdifferentseasons.ThehighestyieldofRNAwasfieldleaves,andthenwaspla2ntlets,barks,fruitsandrootskins.Keywords:Chinesejujube;Organs;Tiss