Stukenberg Daniel, Hensel Tobias, Hoff Josef, Daniel Benjamin, Inckemann René, Tedeschi Jamie N, Nousch Franziska, Fritz Georg
Center for Synthetic Microbiology, Philipps-Universität Marburg, Marburg 35032, Germany.
Max-Planck Institute for Terrestrial Microbiology, Marburg 35043, Germany.
ACS Synth Biol. 2021 Aug 20;10(8):1904-1919. doi: 10.1021/acssynbio.1c00126. Epub 2021 Jul 13.
is known as the world's fastest growing organism with a doubling time of less than 10 min. This incredible growth speed empowers as a chassis for synthetic and molecular biology, potentially replacing in many applications. While first genetic parts have been built and tested for , a comprehensive toolkit containing well-characterized and standardized parts did not exist. To close this gap, we created the Marburg Collection-a highly flexible Golden Gate cloning toolbox optimized for the emerging chassis organism , containing 191 genetic parts. The Marburg Collection overcomes the paradigm of plasmid construction-integrating inserts into a backbone-by enabling the assembly of plasmids from basic genetic parts. This allows users to select the plasmid replication origin and resistance part independently, which is highly advantageous when limited knowledge about the behavior of those parts in the target organism is available. Additional design highlights of the Marburg Collection are novel connector parts, which facilitate modular circuit assembly and, optionally, the inversion of individual transcription units to reduce transcriptional crosstalk in multigene constructs. To quantitatively characterize the genetic parts contained in the Marburg Collection in , we developed a reliable microplate reader measurement workflow for reporter experiments and overcame organism-specific challenges. We think the Marburg Collection with its thoroughly characterized parts will provide a valuable resource for the growing community.
被认为是世界上生长最快的生物体，其倍增时间不到10分钟。这种惊人的生长速度使其成为合成生物学和分子生物学的底盘，在许多应用中可能会取代[原文此处缺失相关内容]。虽然已经构建并测试了[原文此处缺失相关内容]的第一批遗传元件，但尚未存在一个包含经过充分表征和标准化元件的综合工具包。为了填补这一空白，我们创建了马尔堡文库——一个针对新兴底盘生物体[原文此处缺失相关内容]优化的高度灵活的金门克隆工具箱，包含191个遗传元件。马尔堡文库克服了质粒构建的范式——将插入片段整合到骨架中——通过从基本遗传元件中进行质粒的[原文此处缺失相关内容]组装。这允许用户独立选择质粒复制起点和抗性元件，当对这些元件在目标生物体中的行为了解有限时，这具有很大优势。马尔堡文库的其他设计亮点是新型连接元件，它有助于模块化电路组装，并且可以选择反转单个转录单元以减少多基因构建体中的转录串扰。为了定量表征马尔堡文库中包含的[原文此处缺失相关内容]的遗传元件，我们开发了一种用于报告基因实验的可靠微孔板读数器测量工作流程，并克服了特定生物体的挑战。我们认为，具有充分表征元件的马尔堡文库将为不断发展的[原文此处缺失相关内容]社区提供宝贵资源。
