解聚英语

植物叶片经液氮研磨，可使细胞壁破裂，加入去污剂（如CTAB），可使核蛋白体解析，然后使蛋白和多糖杂质沉淀，DNA进入水相，再用酚、氯仿抽提纯化。本实验采用CTAB法，其主要作用是破膜。CTAB是一种非离子去污剂，能溶解膜蛋白与脂肪，也可解聚核蛋白。植物材料在CTAB的处理下，结合65℃水浴使细胞裂解、蛋白质变性、DNA被释放出来。CTAB与核酸形成复合物，此复合物在高盐（>0.7mM）浓度下可溶，并稳定存在，但在低盐浓度（0.1-0.5mMNaCl）下CTAB-核酸复合物就因溶解度降低而沉淀，而大部分的蛋白质及多糖等仍溶解于溶液中。经过氯仿/异戊醇(24:1)抽提去除蛋白质、多糖、色素等来纯化DNA，最后经异丙醇或乙醇等沉淀剂将DNA沉淀分离出来。Plantleavesbygrindinginliquidnitrogen,canmakethecellwallbroken,addingdetergent(suchasCTAB),canmakenuclearproteinanalysis,andthentheproteinandpolysaccharideimpurityprecipitation,DNAintotheaqueousphase,andphenolchloroformextraction,purification.ThisexperimentadoptstheCTABmethod,itsmainroleistoruptureofmembranes.CTABisanonionicdetergent,candissolvethemembraneproteinandfat,canalsobedepolymerizednuclearprotein.PlantmaterialsinCTABtreatment,combinedwith65℃waterbathtocelllysis,proteindenaturation,DNAisreleased.CTABandnucleicacidcomplexes,thecomplexesinhighsalt(>0.7mM)concentrationandsoluble,stable,butatlowsaltconcentration(0.1-0.5mMNaCl)CTAB-nucleicacidcomplexesbecauseofreducedsolubilityandprecipitation,andmostoftheproteinandpolysaccharideisdissolvedinsolution.Bychloroform/isoamylalcohol(24:1)extractiontoremoveproteins,polysaccharides,pigmentsforpurificationofDNA,andfinallybyisopropylalcoholorethanolasprecipitantDNAprecipitationseparated.由于核酸、蛋白质、多糖在特定的紫外波长都有特征吸收。核酸及其衍生物的紫外吸收高峰在260nm。纯的DNA样品A260/280≈1.8，纯的RNA样品A260/280≈2.0，并且1μg/mlDNA溶液A260=0.020。Becausethenucleicacid,protein,polysaccharidehasthecharacteristicabsorptionintheUVwavelengthspecific.NucleicacidanditsderivativeUVabsorptionpeakat260nm.PureDNAsampleswithA260/280≈1.8,pureRNAsampleA260/280≈2,and1μg/mlDNAsolutionA260=0.020