Tian Siqi, Das Rhiju
Department of Biochemistry.
Department of Physics, Stanford University, Stanford, CA 94305, USA.
Bioinformatics. 2017 May 1;33(9):1405-1406. doi: 10.1093/bioinformatics/btw814.
Rapid RNA synthesis of comprehensive single mutant libraries and targeted multiple mutant libraries is enabling new multidimensional chemical approaches to solve RNA structures. PCR assembly of DNA templates and in vitro transcription allow synthesis and purification of hundreds of RNA mutants in a cost-effective manner, with sharing of primers across constructs allowing significant reductions in expense. However, these protocols require organization of primer locations across numerous 96 well plates and guidance for pipetting, non-trivial tasks for which informatics and visualization tools can prevent costly errors. We report here an online tool to accelerate synthesis of large libraries of desired mutants through design and efficient organization of primers. The underlying program and graphical interface have been experimentally tested in our laboratory for RNA domains with lengths up to 300 nucleotides and libraries encompassing up to 960 variants. In addition to the freely available Primerize-2D server, the primer design code is available as a stand-alone Python package for broader applications.
http://primerize2d.stanford.edu.
Supplementary data are available at Bioinformatics online.
全面的单突变体文库和靶向多突变体文库的快速RNA合成，正在催生新的多维化学方法来解析RNA结构。DNA模板的PCR组装和体外转录能够以经济高效的方式合成和纯化数百个RNA突变体，通过跨构建体共享引物可显著降低成本。然而，这些方案需要在众多96孔板中安排引物位置并提供移液指导，这是复杂的任务，信息学和可视化工具可防止出现代价高昂的错误。我们在此报告一种在线工具，通过设计和高效组织引物来加速所需突变体大文库的合成。基础程序和图形界面已在我们实验室针对长度达300个核苷酸的RNA结构域以及包含多达960个变体的文库进行了实验测试。除了免费提供的Primerize-2D服务器外，引物设计代码还作为独立的Python包提供，以供更广泛的应用。
http://primerize2d.stanford.edu。
补充数据可在《生物信息学》在线获取。
