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A ubiquitous amino acid source for prokaryotic and eukaryotic cell-free transcription-translation systems

Nagappa, Lakshmeesha N, Sato, Wakana, Alam, Farzana, Chengan, Kameshwari, Smales, Christopher Mark, von der Haar, Tobias, Polizzi, Karen M., Adamala, Katarzyna, Moore, Simon J. (2022) A ubiquitous amino acid source for prokaryotic and eukaryotic cell-free transcription-translation systems. Frontiers in Bioengineering and Biotechnology, 10 . Article Number 992708. ISSN 2296-4185. (doi:10.3389/fbioe.2022.992708) (KAR id:102961)


Cell-free gene expression (CFE) systems are an attractive tool for engineering within synthetic biology and for industrial production of high-value recombinant proteins. CFE reactions require a cell extract, energy system, amino acids, and DNA, to catalyse mRNA transcription and protein synthesis. To provide an amino acid source, CFE systems typically use a commercial standard, which is often proprietary. Herein we show that a range of common microbiology rich media (i.e., tryptone, peptone, yeast extract and casamino acids) unexpectedly provide an effective and low-cost amino acid source. We show that this approach is generalisable, by comparing batch variability and protein production in the following range of CFE systems: Escherichia coli (Rosetta™ 2 (DE3), BL21(DE3)), Streptomyces venezuelae and Pichia pastoris. In all CFE systems, we show equivalent or increased protein synthesis capacity upon replacement of the commercial amino acid source. In conclusion, we suggest rich microbiology media provides a new amino acid source for CFE systems with potential broad use in synthetic biology and industrial biotechnology applications.

Item Type: Article
DOI/Identification number: 10.3389/fbioe.2022.992708
Additional information: For the purpose of open access, the author has applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission.
Subjects: Q Science > QR Microbiology
Divisions: Divisions > Division of Natural Sciences > Biosciences
Funders: Leverhulme Trust (
National Science Foundation (
Engineering and Physical Sciences Research Council (
Depositing User: Tobias von der Haar
Date Deposited: 26 Sep 2023 12:19 UTC
Last Modified: 27 Feb 2024 11:19 UTC
Resource URI: (The current URI for this page, for reference purposes)

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