StreptoCAD:一个开源软件工具箱,自动化链霉菌基因组工程工作流程。

IF 3.9 2区 生物学 Q1 BIOCHEMICAL RESEARCH METHODS
Lucas Levassor, Christopher M Whitford, Søren D Petersen, Kai Blin, Tilmann Weber, Rasmus J N Frandsen
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引用次数: 0

摘要

链霉菌在发现用于医药或可持续农业的新型生物活性分子方面具有巨大的潜力。然而,目前的链霉菌基因工程方法涉及为每个目标基因手工设计复杂的实验分子生物工程策略,这阻碍了高通量的探索。在这里,我们介绍StreptoCAD,一个开源软件工具箱,自动化和简化链霉菌基因组工程策略的设计,支持各种基于crispr和基因过表达的方法。一旦启动,StreptoCAD设计所有必要的DNA引物和CRISPR引导序列,模拟质粒组装(克隆)和由此产生的基因组靶标修饰,并进一步总结实验室实施和文档所需的信息。StreptoCAD目前提供六种设计工作流程,包括构建过表达文库、碱基编辑(包括多路CRISPR-BEST质粒生成)以及使用CRISPR-Cas9、CRISPR-Cas3和CRISPRi系统进行基因组工程。除了自动化设计过程外,StreptoCAD还进一步确保符合FAIR原则,通过标准化输出文件确保再现性和易于数据管理。为了实验证明StreptoCAD的设计过程和输出,我们设计并构建了一系列基因过表达菌株,并在Streptomyces Gö40/10中进行了CRISPRi敲低,强调了该工具的效率和用户友好性。该工具简化了复杂的基因工程任务,并通过标准化的工作流程和设计参数促进了协作。StreptoCAD将改变链霉菌的基因组工程,使所有人都能进行复杂的基因操作,并加速天然产物的发现。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
StreptoCAD: An Open-Source Software Toolbox Automating Genome Engineering Workflows in Streptomycetes.

Streptomycetes hold immense potential for discovering novel bioactive molecules for applications in medicine or sustainable agriculture. However, high-throughput exploration is hampered by the current Streptomyces genetic engineering methods that involve the manual design of complex experimental molecular biological engineering strategies for each targeted gene. Here, we introduce StreptoCAD, an open-source software toolbox that automates and streamlines the design of genome engineering strategies in Streptomyces, supporting various CRISPR-based and gene overexpression methods. Once initiated, StreptoCAD designs all necessary DNA primers and CRISPR guide sequences, simulates plasmid assemblies (cloning) and the resulting modification of the genomic target(s), and further summarizes the information needed for laboratory implementation and documentation. StreptoCAD currently offers six design workflows, including the construction of overexpression libraries, base-editing, including multiplexed CRISPR-BEST plasmid generation, and genome engineering using CRISPR-Cas9, CRISPR-Cas3, and CRISPRi systems. In addition to automating the design process, StreptoCAD further secures compliance with the FAIR principles, ensuring reproducibility and ease of data management via standardized output files. To experimentally demonstrate the design process and output of StreptoCAD, we designed and constructed a series of gene overexpression strains, and performed CRISPRi knockdowns in Streptomyces Gö40/10, underscoring the tool's efficiency and user-friendliness.. This tool simplifies complex genetic engineering tasks and promotes collaboration through standardized workflows and design parameters. StreptoCAD is set to transform genome engineering in Streptomyces, making sophisticated genetic manipulations accessible for all and accelerating natural product discovery.

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来源期刊
CiteScore
8.00
自引率
10.60%
发文量
380
审稿时长
6-12 weeks
期刊介绍: The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism. Topics may include, but are not limited to: Design and optimization of genetic systems Genetic circuit design and their principles for their organization into programs Computational methods to aid the design of genetic systems Experimental methods to quantify genetic parts, circuits, and metabolic fluxes Genetic parts libraries: their creation, analysis, and ontological representation Protein engineering including computational design Metabolic engineering and cellular manufacturing, including biomass conversion Natural product access, engineering, and production Creative and innovative applications of cellular programming Medical applications, tissue engineering, and the programming of therapeutic cells Minimal cell design and construction Genomics and genome replacement strategies Viral engineering Automated and robotic assembly platforms for synthetic biology DNA synthesis methodologies Metagenomics and synthetic metagenomic analysis Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction Gene optimization Methods for genome-scale measurements of transcription and metabolomics Systems biology and methods to integrate multiple data sources in vitro and cell-free synthetic biology and molecular programming Nucleic acid engineering.
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