The Marburg Collection: A Golden Gate DNA Assembly Framework for Synthetic Biology Applications in Vibrio natriegens

Daniel Stukenberg, Tobias Hensel, Josef Hoff, Benjamin Daniel, René Inckemann, Jamie N. Tedeschi, Franziska Nousch, Georg Fritz

Research output: Contribution to journalArticlepeer-review

5 Citations (Web of Science)

Abstract

Vibrio natriegens is known as the world's fastest growing organism with a doubling time of less than 10 min. This incredible growth speed empowers V. natriegens as a chassis for synthetic and molecular biology, potentially replacing E. coli in many applications. While first genetic parts have been built and tested for V. natriegens, 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 V. natriegens, containing 191 genetic parts. The Marburg Collection overcomes the paradigm of plasmid construction-integrating inserts into a backbone-by enabling the de novo 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 V. natriegens, 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 V. natriegens community.

Original languageEnglish
Pages (from-to)1904-1919
JournalACS Synthetic Biology
Volume10
Issue number8
Early online date13 Jul 2021
DOIs
Publication statusPublished - 20 Aug 2021

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