Genetic circuit performance under conditions relevant for industrial bioreactors

Felix Moser, Nicolette J. Broers, Sybe Hartmans, Alvin Tamsir, Richard Kerkman, J. Roubos, Roel Bovenberg, Christopher A. Voigt

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Synthetic genetic programs promise to enable novel applications in industrial processes. For such applications, the genetic circuits that compose programs will require fidelity in varying and complex environments. In this work, we report the performance of two synthetic circuits in Escherichia coli under industrially relevant conditions, including the selection of media, strain, and growth rate. We test and compare two transcriptional circuits: an AND and a NOR gate. In E. coli DH10B, the AND gate is inactive in minimal media; activity can be rescued by supplementing the media and transferring the gate into the industrial strain E. coli DS68637 where normal function is observed in minimal media. In contrast, the NOR gate is robust to media composition and functions similarly in both strains. The AND gate is evaluated at three stages of early scale-up: 100 mL shake flask experiments, a 1 mL MTP microreactor, and a 10 L bioreactor. A reference plasmid that constitutively produces a GFP reporter is used to make comparisons of circuit performance across conditions. The AND gate function is quantitatively different at each scale. The output deteriorates late in fermentation after the shift from exponential to constant feed rates, which induces rapid resource depletion and changes in growth rate. In addition, one of the output states of the AND gate failed in the bioreactor, effectively making it only responsive to a single input. Finally, cells carrying the AND gate show considerably less accumulation of biomass. Overall, these results highlight challenges and suggest modified strategies for developing and characterizing genetic circuits that function reliably during fermentation.

Original languageEnglish
Pages (from-to)555-564
Number of pages10
JournalACS Synthetic Biology
Volume1
Issue number11
DOIs
Publication statusPublished - Nov 16 2012

Fingerprint

Bioreactors
Escherichia coli
Fermentation
Networks (circuits)
Growth
Biomass
Plasmids
Strain rate
Chemical analysis
Experiments

Keywords

  • computer-aided design
  • fermentation
  • genetic compiler
  • RBS calculator
  • synthetic biology
  • systems biology

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology (miscellaneous)
  • Biomedical Engineering

Cite this

Moser, F., Broers, N. J., Hartmans, S., Tamsir, A., Kerkman, R., Roubos, J., ... Voigt, C. A. (2012). Genetic circuit performance under conditions relevant for industrial bioreactors. ACS Synthetic Biology, 1(11), 555-564. https://doi.org/10.1021/sb3000832

Genetic circuit performance under conditions relevant for industrial bioreactors. / Moser, Felix; Broers, Nicolette J.; Hartmans, Sybe; Tamsir, Alvin; Kerkman, Richard; Roubos, J.; Bovenberg, Roel; Voigt, Christopher A.

In: ACS Synthetic Biology, Vol. 1, No. 11, 16.11.2012, p. 555-564.

Research output: Contribution to journalArticle

Moser, F, Broers, NJ, Hartmans, S, Tamsir, A, Kerkman, R, Roubos, J, Bovenberg, R & Voigt, CA 2012, 'Genetic circuit performance under conditions relevant for industrial bioreactors', ACS Synthetic Biology, vol. 1, no. 11, pp. 555-564. https://doi.org/10.1021/sb3000832
Moser, Felix ; Broers, Nicolette J. ; Hartmans, Sybe ; Tamsir, Alvin ; Kerkman, Richard ; Roubos, J. ; Bovenberg, Roel ; Voigt, Christopher A. / Genetic circuit performance under conditions relevant for industrial bioreactors. In: ACS Synthetic Biology. 2012 ; Vol. 1, No. 11. pp. 555-564.
@article{e48c12c81bc04da6a9f863ee42078e42,
title = "Genetic circuit performance under conditions relevant for industrial bioreactors",
abstract = "Synthetic genetic programs promise to enable novel applications in industrial processes. For such applications, the genetic circuits that compose programs will require fidelity in varying and complex environments. In this work, we report the performance of two synthetic circuits in Escherichia coli under industrially relevant conditions, including the selection of media, strain, and growth rate. We test and compare two transcriptional circuits: an AND and a NOR gate. In E. coli DH10B, the AND gate is inactive in minimal media; activity can be rescued by supplementing the media and transferring the gate into the industrial strain E. coli DS68637 where normal function is observed in minimal media. In contrast, the NOR gate is robust to media composition and functions similarly in both strains. The AND gate is evaluated at three stages of early scale-up: 100 mL shake flask experiments, a 1 mL MTP microreactor, and a 10 L bioreactor. A reference plasmid that constitutively produces a GFP reporter is used to make comparisons of circuit performance across conditions. The AND gate function is quantitatively different at each scale. The output deteriorates late in fermentation after the shift from exponential to constant feed rates, which induces rapid resource depletion and changes in growth rate. In addition, one of the output states of the AND gate failed in the bioreactor, effectively making it only responsive to a single input. Finally, cells carrying the AND gate show considerably less accumulation of biomass. Overall, these results highlight challenges and suggest modified strategies for developing and characterizing genetic circuits that function reliably during fermentation.",
keywords = "computer-aided design, fermentation, genetic compiler, RBS calculator, synthetic biology, systems biology",
author = "Felix Moser and Broers, {Nicolette J.} and Sybe Hartmans and Alvin Tamsir and Richard Kerkman and J. Roubos and Roel Bovenberg and Voigt, {Christopher A.}",
year = "2012",
month = "11",
day = "16",
doi = "10.1021/sb3000832",
language = "English",
volume = "1",
pages = "555--564",
journal = "ACS Synthetic Biology",
issn = "2161-5063",
publisher = "American Chemical Society",
number = "11",

}

TY - JOUR

T1 - Genetic circuit performance under conditions relevant for industrial bioreactors

AU - Moser, Felix

AU - Broers, Nicolette J.

AU - Hartmans, Sybe

AU - Tamsir, Alvin

AU - Kerkman, Richard

AU - Roubos, J.

AU - Bovenberg, Roel

AU - Voigt, Christopher A.

PY - 2012/11/16

Y1 - 2012/11/16

N2 - Synthetic genetic programs promise to enable novel applications in industrial processes. For such applications, the genetic circuits that compose programs will require fidelity in varying and complex environments. In this work, we report the performance of two synthetic circuits in Escherichia coli under industrially relevant conditions, including the selection of media, strain, and growth rate. We test and compare two transcriptional circuits: an AND and a NOR gate. In E. coli DH10B, the AND gate is inactive in minimal media; activity can be rescued by supplementing the media and transferring the gate into the industrial strain E. coli DS68637 where normal function is observed in minimal media. In contrast, the NOR gate is robust to media composition and functions similarly in both strains. The AND gate is evaluated at three stages of early scale-up: 100 mL shake flask experiments, a 1 mL MTP microreactor, and a 10 L bioreactor. A reference plasmid that constitutively produces a GFP reporter is used to make comparisons of circuit performance across conditions. The AND gate function is quantitatively different at each scale. The output deteriorates late in fermentation after the shift from exponential to constant feed rates, which induces rapid resource depletion and changes in growth rate. In addition, one of the output states of the AND gate failed in the bioreactor, effectively making it only responsive to a single input. Finally, cells carrying the AND gate show considerably less accumulation of biomass. Overall, these results highlight challenges and suggest modified strategies for developing and characterizing genetic circuits that function reliably during fermentation.

AB - Synthetic genetic programs promise to enable novel applications in industrial processes. For such applications, the genetic circuits that compose programs will require fidelity in varying and complex environments. In this work, we report the performance of two synthetic circuits in Escherichia coli under industrially relevant conditions, including the selection of media, strain, and growth rate. We test and compare two transcriptional circuits: an AND and a NOR gate. In E. coli DH10B, the AND gate is inactive in minimal media; activity can be rescued by supplementing the media and transferring the gate into the industrial strain E. coli DS68637 where normal function is observed in minimal media. In contrast, the NOR gate is robust to media composition and functions similarly in both strains. The AND gate is evaluated at three stages of early scale-up: 100 mL shake flask experiments, a 1 mL MTP microreactor, and a 10 L bioreactor. A reference plasmid that constitutively produces a GFP reporter is used to make comparisons of circuit performance across conditions. The AND gate function is quantitatively different at each scale. The output deteriorates late in fermentation after the shift from exponential to constant feed rates, which induces rapid resource depletion and changes in growth rate. In addition, one of the output states of the AND gate failed in the bioreactor, effectively making it only responsive to a single input. Finally, cells carrying the AND gate show considerably less accumulation of biomass. Overall, these results highlight challenges and suggest modified strategies for developing and characterizing genetic circuits that function reliably during fermentation.

KW - computer-aided design

KW - fermentation

KW - genetic compiler

KW - RBS calculator

KW - synthetic biology

KW - systems biology

UR - http://www.scopus.com/inward/record.url?scp=84869451704&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84869451704&partnerID=8YFLogxK

U2 - 10.1021/sb3000832

DO - 10.1021/sb3000832

M3 - Article

VL - 1

SP - 555

EP - 564

JO - ACS Synthetic Biology

JF - ACS Synthetic Biology

SN - 2161-5063

IS - 11

ER -