Horsley Nicole, von Barnau Sythoff Adam, Delgado Mark, Liu Selina, Cabernard Clemens
Department of Biology, University of Washington, Life Science Building, Seattle WA 98195, USA.
Current address: Department of Stem Cell Biology and Regenerative Medicine, Stanford University, 265 Campus Drive, Stanford CA 94305.
bioRxiv. 2025 Mar 2:2025.02.28.640916. doi: 10.1101/2025.02.28.640916.
CRISPR/Cas9 is a powerful tool for targeted genome engineering experiments. With CRISPR/Cas9, genes can be deleted or modified by inserting small peptides, fluorescent proteins or other tags for protein labelling experiments. Such experiments are important for detailed protein characterization . However, designing and cloning the corresponding constructs can be repetitive, time consuming and laborious. To aid users in CRISPR/Cas9-based genome engineering experiments, we built CrisprBuildr, a web-based application that allows users to delete genes or insert fluorescent proteins at the N- or C-terminus of their gene of choice. The application is built on the genome but can be used as a template for other available genomes. We have also generated new tagging vectors, using EGFP and mCherry combined with the small peptide SspB-Q73R for use in iLID-based optogenetic experiments. CrisprBuildr guides users through the process of designing guide RNAs and repair template vectors. CrisprBuildr is an open-source application and future releases could incorporate additional tagging or deletion vectors, genomes or CRISPR applications.
CRISPR/Cas9是用于靶向基因组工程实验的强大工具。利用CRISPR/Cas9，可以通过插入小肽、荧光蛋白或其他用于蛋白质标记实验的标签来删除或修饰基因。此类实验对于详细的蛋白质表征很重要。然而，设计和克隆相应的构建体可能是重复性的、耗时的且费力的。为了帮助用户进行基于CRISPR/Cas9的基因组工程实验，我们构建了CrisprBuildr，这是一个基于网络的应用程序，允许用户在其选择的基因的N端或C端删除基因或插入荧光蛋白。该应用程序基于该基因组构建，但可作为其他可用基因组的模板。我们还生成了新的标签载体，使用EGFP和mCherry与小肽SspB-Q73R组合，用于基于iLID的光遗传学实验。CrisprBuildr指导用户完成设计引导RNA和修复模板载体的过程。CrisprBuildr是一个开源应用程序，未来版本可能会纳入更多的标签或删除载体、基因组或CRISPR应用。
