Laboratory of Biomolecular Research, Paul Scherrer Institut, Villigen, Switzerland and Department of Biology, ETH Zurich, Zurich, Switzerland.
PLoS One. 2013 Oct 30;8(10):e78878. doi: 10.1371/journal.pone.0078878. eCollection 2013.
Scanning mutagenesis is a powerful protein engineering technique used to study protein structure-function relationship, map binding sites and design more stable proteins or proteins with altered properties. One of the time-consuming tasks encountered in application of this technique is the design of primers for site-directed mutagenesis. Here we present an open-source multi-platform software AAscan developed to design primers for this task according to a set of empirical rules such as melting temperature, overall length, length of overlap regions, and presence of GC clamps at the 3' end, for any desired substitution. We also describe additional software tools which are used to analyse a large number of sequencing results for the presence of desired mutations, as well as related software to design primers for ligation independent cloning. We have used AAscan software to design primers to make over 700 mutants, with a success rate of over 80%. We hope that the open-source nature of our software and ready availability of freeware tools used for its development will facilitate its adaptation and further development. The software is distributed under GPLv3 licence and is available at http://www.psi.ch/lbr/aascan.
扫描诱变是一种强大的蛋白质工程技术，用于研究蛋白质结构-功能关系、绘制结合位点和设计更稳定或具有改变性质的蛋白质。在应用该技术时遇到的耗时任务之一是设计定点诱变的引物。我们在此介绍一种开源的多平台软件 AAscan，该软件是根据一套经验规则（如熔点、全长、重叠区长度和 3'端的 GC 夹）设计引物的，可用于任何所需的取代。我们还描述了其他用于分析大量测序结果以确定所需突变存在的软件工具，以及用于连接独立克隆的引物设计相关软件。我们使用 AAscan 软件设计了超过 700 个突变体的引物，成功率超过 80%。我们希望我们的软件的开源性质和用于其开发的免费软件工具的可用性将促进其适应和进一步发展。该软件根据 GPLv3 许可证分发，可在 http://www.psi.ch/lbr/aascan 上获得。
