4 PCR技術.ppt

PCR技术;内容;Polymerase chain reaction， 聚合酶链式反应;; PCR技术操作简便，可以在试管中建立反应，经数小时之后，就能将极微量的目的基因或某一特定的DNA片段扩增109倍，且无需烦琐的基因克隆程序，便可获得足够数量的精确的DNA拷贝。;PCR扩增过程;;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty Ltd.;* 30次循环后靶序列扩增的数量;理论上为2n。 30次循环，DNA产量达109拷贝 实际上为(1+R)n。R为扩增效率，平均为75% 30次循环，DNA产量实际为106-107拷贝;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty Ltd.;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty Ltd.;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty Ltd.;;第二节 PCR反应体系中的各组分分析;;;引物的保存和使用浓度;已知A基因的cDNA序列如下，起始密码和终止密码均用框标出，试设计一对引物扩增该基因的ORF。 TTCTAAAAAAGAAGAAAAATGTCCGAAGTAATCGAAGAACATCTTCTCAGCGATAATTCTGATGATTCCAGCTCGGAATTGACTTCTACAGAGGAAAATTGGGAAGAAATTTTTGCAGATTTTCAAAGCTGTCAGGAGGATTCTGTTGTGGGAACCCCGCCAGAAGCGGCGGCCGGAGGAGGTTGTTCGAAGGATTGGAACCGGTATAAGGGCGTTAGACGGCGGCCGTGGGGGAAGTTCGCGGCGGATTCGAATATTTCCGGACCCGTTAGAGTAAACCCGAGAAAACGTTTCCCTGCGGAGCCTTCTACGACGTCGTCGTCTTCTTCTTCTTCTTCGTCTGAAAATAGTGGAGGAAGGACGACGAGACGATCTTAATTAATAGGAGTTAAGCCCGATAATCCCTCATTGTAAAATTATTAACTTCATCGATGTTCTTTGGAATGTACAAAATTTTATATCCAAAAAAGTTCACAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA;;;二、DNA Pol;Taq DNA pol的发现;用于PCR的pol种类;各种耐热DNA pol的特性;Taq DNA pol的灭活;Taq DNA pol的使用浓度;三、DNA 模板;DNA 模板的用量;DNA 模板的纯度;DNA模板的制备;四、dNTPs;五、PCR反应的缓冲液;缓冲液配方;六、Mg2+浓度;七、PCR反应条件的选择;变性温度与时间;退火温度与时间;延伸温度与时间;延伸终止;循环次数;八、标准的PCR反应体系;;;第三节 PCR技术的分类;正常PCR（Normal PCR）;巢式PCR（Nested PCR）;;分类;半巢式PCR;中途进退式PCR;巢式PCR的应用;复合PCR;;复合PCR产物的电泳图谱;复合PCR的应用;复合PCR的应用;复合PCR的应用;;多重PCR的特点;不对称PCR;;不对称PCR的应用;反转录PCR;;RT-PCR的应用;One-step RT-PCR;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty Ltd.;RT-PCR;定量PCR;何为实时？;SYBR?Green?I荧光染料;起始模版浓度定量;荧光定量PCR工作曲线;Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2011 Aspose Pty