# Development of a Bacillus subtilis cell-free transcription-translation system for prototyping regulatory elements

Article · Richard J. R. Kelwick, Alexander J. Webb, James T. MacDonald, Paul S. Freemont

- Archive id: doi:10.1016/j.ymben.2016.09.008
- Published: 2016-10-06
- Venue: Metabolic Engineering
- DOI: https://doi.org/10.1016/j.ymben.2016.09.008
- PMID: 27697563
- Citations: 135
- Source URL: https://doi.org/10.1016/j.ymben.2016.09.008
- Provenance: openalex

## Abstract

Cell-free transcription-translation systems were originally applied towards in vitro protein production. More recently, synthetic biology is enabling these systems to be used within a systematic design context for prototyping DNA regulatory elements, genetic logic circuits and biosynthetic pathways . The Gram-positive soil bacterium , Bacillus subtilis , is an established model organism of industrial importance. To this end, we developed several B. subtilis -based cell-free systems. Our improved B. subtilis WB800N-based system was capable of producing 0.8 µM GFP, which gave a ~72x fold-improvement when compared with a B. subtilis 168 cell-free system. Our improved system was applied towards the prototyping of a B. subtilis promoter library in which we engineered several promoters, derived from the wild-type P grac ( σ A) promoter, that display a range of comparable in vitro and in vivo transcriptional activities. Additionally, we demonstrate the cell-free characterisation of an inducible expression system , and the activity of a model enzyme - renilla luciferase .

_Abstract reproduced under CC-BY (publisher license, via OpenAlex)._

Topics: [Synthetic Biology & Genome Engineering](https://space-kitty.com/topics/synthetic-biology)

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