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聚丙烯酰胺凝胶电泳.doc

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聚丙烯酰胺凝胶电泳凝胶的配置表1配制Tris-甘氨酸SDS-PAGE聚丙烯酰胺凝胶电泳分离胶所用溶液溶液成分不同体积（ml）凝胶液中各成分所需体积（ml）5101520253040508%水2.34.66.99.311.513.918.523.230%丙烯酰胺溶液1.32.745.36.7810.713.31.5mol/LTris(pH8.8)1.32.53.856.37.51012.510%SDS0.050.10.150.20.250.30.40.510%过硫酸铵0.050.10.150.20.250.30.40.5TEMED0.0030.0060.0090.0120.0150.0180.0240.0310%水1.945.97.99.911.915.919.830%丙烯酰胺溶液1.73.356.78.31013.316.71.5mol/LTris(pH8.8)1.32.53.856.37.51012.510%SDS0.050.10.150.20.250.30.40.510%过硫酸铵0.050.10.150.20.250.30.40.5TEMED0.0020.0040.0060.0080.010.0120.0160.0212%水1.63.34.96.68.29.913.216.530%丙烯酰胺溶液2468101216201.5mol/LTris(pH8.8)1.32.53.856.37.51012.510%SDS0.050.10.150.20.250.30.40.510%过硫酸铵0.050.10.150.20.250.30.40.5TEMED0.0020.0040.0060.0080.010.0120.0160.0212.5%水1.53.14.656.37.89.412.515.6630%丙烯酰胺溶液2.14.26.258.310.412.516.720.841.5mol/LTris(pH8.8)1.32.53.856.37.51012.510%SDS0.050.10.150.20.250.30.40.510%过硫酸铵0.050.10.150.20.250.30.40.5TEMED0.0020.0040.0060.0080.010.0120.0160.0215%水1.12.33.44.65.76.99.211.530%丙烯酰胺溶液2.557.51012.51520251.5mol/LTris(pH8.8)1.32.53.856.37.51012.510%SDS0.050.10.150.20.250.30.40.510%过硫酸铵0.050.10.150.20.250.30.40.5TEMED0.0020.0040.0060.0080.010.0120.0160.02表2配制Tris-甘氨酸SDS-PAGE聚丙烯酰胺凝胶电泳5%积层胶所用溶液溶液成分不同体积（ml）凝胶液中各成分所需体积（ml）1234568105%水0.681.42.12.73.44.15.56.830%丙烯酰胺溶液0.170.330.50.670.8311.31.71.0mol/LTris(pH6.8)0.130.250.380.50.630.7501.2510%SDS0.010.020.030.040.050.060.080.110%过硫酸铵0.010.020.030.040.050.060.080.1TEMED0.0010.0020.0030.0040.0050.0060.0080.01相关溶液贮液的配制30%聚丙烯酰胺贮液：丙烯酰胺150g甲叉双丙烯酰胺4gMilliQ水500ml滤纸过滤后，棕色瓶4℃冰箱保存。1.5mol/LTris-basepH8.8：Tris碱90.75gMilliQ水400ml用1mol/LHCl调pH至8.8，加MilliQ水定容至500ml4℃冰箱保存。1.0mol/LTris-basepH6.8:Tris-base12.11gMilliQ水80ml用1mol/LHCl调pH至6.8，加MilliQ水定容至100ml4℃冰箱保存。10%SDS：SDS10gMilliQ水100ml混匀后，室温保存。10%Ap：Ap0.1gMilliQ水1ml（用时加水溶解）溶解后，4℃冰箱保存。10×电泳缓冲液（1×=25mMTris，192mMglycine，0.1%SDS，pH8.3）：Tris碱30g甘氨酸144gSDS10gMilliQ水1L混匀后，室温保存。Loadingbuffer2×4×1.0MTris-HClPH6.81ml2ml甘油2ml4ml10%(w/v)SDS4mlSDS0.8g0.25%(w/v)溴酚蓝0.2ml0.4mlβ-巯基乙醇1ml2mlmilliQ水定容至10ml10ml5×SDS-PAGELoadingB