[第四章 酶分子修饰.pptVIP

4.6.4 酶分子定向进化的选择策略 定向进化中，突变具有随机性，但通过选择特定方向的突变限定了进化趋势，加之控制实验条件，限定突变种类， 降低突变率，缩小突变库的容量，这不仅减少了工作量，更重要的是加快了酶在某一方向的进化速度。 筛选方法必须灵敏，至少与目的性质相关。另有一些其他的筛选方法，如加入能产生可见光信号的底物或利用绿色荧光蛋白的荧光性质等。 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. 1、突变基因文库的构建 定义：将各种不同突变基因与载体重组，再转入适宜的细胞或包装成重组λ噬菌体的技术过程。 质量要求：文库包容性和文库完整性 过程： 载体选择 基因重组 形成基因文库 质粒 噬菌体DNA 黏粒载体 噬菌粒载体 转化 转导 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. 2、突变基因的筛选 （1）定向选择环境条件的设定 （2）高通量筛选技术 平板筛选技术 荧光筛选法 噬菌体表面展示法 酵母细胞表面展示法 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. 举例 例1：定向进化提高枯草芽孢杆菌脂肪酶的活力 例2：大肠杆菌碱性磷酸酶的体外定向进化研究 例3：采用易错PCR对粘质沙雷氏菌几丁质酶C进行定向进化 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. 上节课主要内容回顾 从酶的生产到酶的改性 酶分子修饰 所要解决的问题和修饰的目标 酶分子修饰的种类 侧链基团修饰 Evaluation only. Created with Aspose.Slides for .NET 3.5 Client Profile 5.2.0.0. Copyright 2004-2