中南大学 生物化学 考研课件 ch12 核酸结构 structure of nucleic acids

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1、Chapter12Structureofnucleicacids核酸结构ContentsDNAstructureThethreelevelsofDNAstructureRNAstructure(self-study自学)DifferenceinstructureofmRNAbetweenprokaryoticcellsandeucaryoticcellsSecondaryandtertiarystructureofRNAThethreelevelsofDNAstructure（DNA的三级结构）Theprima

2、rystructure一级结构DNASequencing测序Thesecondarystructure二级结构Basicknowledge基本知识ComparisonamongA,B,Z-conformationformsofDNA(DNA的A,B,Z构象形式的比较）DenaturationandrenaturationofDNA（DNA）变性与复性OtherformsofDNAconformation(DNA构象的其它形式)Thetertiarystructure三级结构DNAorganizationinch

3、romosomes染色体中的DNA构架TheprimarystructureofDNA（DNA一级结构）1ststructureofDNAisDNAsequence(basesequence)是DNA序列或碱基序列DNAsequencing（DNA测序）MethodsChemicalmethod,alsocalledMaxam-Gilbertmethod(1977)化学法Chainterminationmethod,alsocalledSangerdideoxymethod链终止法或Sanger双脱氧方法no

4、wthemethodofchoice常用方法.PrincipleofChainterminationmethodorSangerdideoxymethod链终止法或Sanger双脱氧方法原理WhenaddNTPsisaddedtotheextendingDNAchain,thenthechainextensionwillbeterminatedduetothenearlyaddedddNTPslacks3’-OH.如果有某一个双脱氧核苷酸连接在延伸的DNA链上，那么，由于新连接上的该双脱氧核苷酸缺乏DNA链延伸

5、所需要的3’-OH，DNA链延伸终止。WhenthefourkindsofddNTPs(ddATP,ddGTP,ddTTP,ddCTP)bindtotheextendingDNAatanytimeduringsynthesisofDNAaccordingtobasepairprinciple,thenaseriesofDNApiecescanbesynthesized,analysisofthesizeoftheseDNApiecescangivethesequenceoftheDNA.当四种双脱氧核苷酸在基于

6、碱基配对原则进行链延伸过程中，随机键合到新延伸的DNA链上，那么，便能形成一系列长短不一的DNA链，分析这些链的分子量大小，会给出整个DNA链碱基序列信息。ReviewonthestructuresofATP,dATP,ddATPDNAsequencing（DNA测序）ChainterminationmethodDNAsequencing（DNA测序）ChainterminationmethodDNAsynthesisMethodologyofChainterminationmethodorSangerdide

7、oxymethod链终止法或Sanger双脱氧方法的方法论ManuelmethodAutoradiography放射自显影法AutomaticmethodFluorescentdyes荧光染料（显影）ExamplesofgelsforsequencingofDNA测序用胶示例ChainterminationmethodI---autoradiography放射自显影法Incubationmixtureinfourdifferenttubes在四个试管中分别进行DNA链延伸实验Single-strandedDNA

8、templatePrimer引物DNApolymeraseI(DNA聚合酶I)dNTPs,oneofwhichisradioactivelylabeled,e.g.,(a-32P)dATPddATP,ddTTP,ddGTP,orddCTPElectrophoresis电泳Membranetransferandauto-radiophotography转膜和放射自显影Analysisof