Abstract
Synthetic biology is rapidly evolving into a new phase that emphasizes real-world applications such as environmental remediation. Recently, Comamonas testosteroni has become a promising chassis for bioremediation due to its natural pollutant-degrading capacity; however, its application is hindered by the lack of fundamental gene expression tools. Here, we present a synthetic biology toolkit that enables rapid creation of functional gene circuits in C. testosteroni. We first built a shuttle system that allows efficient circuit construction in E. coli and necessary phenotypic testing in C. testosteroni. Then, we tested a set of wildtype inducible promoters, and further used a hybrid strategy to create engineered promoters to expand expression strength and dynamics. Additionally, we tested the T7 RNA Polymerase-PT7 promoter system and reduced its leaky expression through promoter mutation for gene expression. By coupling random library construction with FACS screening, we further developed a synthetic T7 promoter library to confer a wider range of expression strength and dynamic characteristics. This study provides a set of valuable tools to engineer gene circuits in C. testosteroni, facilitating the establishment of the organism as a useful microbial chassis for bioremediation purposes.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1753-1762 |
| Number of pages | 10 |
| Journal | ACS synthetic biology |
| Volume | 7 |
| Issue number | 7 |
| DOIs | |
| State | Published - Jul 20 2018 |
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Keywords
- Comamonas testosteroni
- P promoter library
- T7 RNA polymerase-P system
- inducible promoter
- shuttle vehicle
- synthetic biology toolkit
ASJC Scopus subject areas
- Biomedical Engineering
- Biochemistry, Genetics and Molecular Biology (miscellaneous)
Cite this
Developing a Synthetic Biology Toolkit for Comamonas testosteroni, an Emerging Cellular Chassis for Bioremediation. / Tang, Qiang; Lu, Ting; Liu, Shuang Jiang.
In: ACS synthetic biology, Vol. 7, No. 7, 20.07.2018, p. 1753-1762.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Developing a Synthetic Biology Toolkit for Comamonas testosteroni, an Emerging Cellular Chassis for Bioremediation
AU - Tang, Qiang
AU - Lu, Ting
AU - Liu, Shuang Jiang
PY - 2018/7/20
Y1 - 2018/7/20
N2 - Synthetic biology is rapidly evolving into a new phase that emphasizes real-world applications such as environmental remediation. Recently, Comamonas testosteroni has become a promising chassis for bioremediation due to its natural pollutant-degrading capacity; however, its application is hindered by the lack of fundamental gene expression tools. Here, we present a synthetic biology toolkit that enables rapid creation of functional gene circuits in C. testosteroni. We first built a shuttle system that allows efficient circuit construction in E. coli and necessary phenotypic testing in C. testosteroni. Then, we tested a set of wildtype inducible promoters, and further used a hybrid strategy to create engineered promoters to expand expression strength and dynamics. Additionally, we tested the T7 RNA Polymerase-PT7 promoter system and reduced its leaky expression through promoter mutation for gene expression. By coupling random library construction with FACS screening, we further developed a synthetic T7 promoter library to confer a wider range of expression strength and dynamic characteristics. This study provides a set of valuable tools to engineer gene circuits in C. testosteroni, facilitating the establishment of the organism as a useful microbial chassis for bioremediation purposes.
AB - Synthetic biology is rapidly evolving into a new phase that emphasizes real-world applications such as environmental remediation. Recently, Comamonas testosteroni has become a promising chassis for bioremediation due to its natural pollutant-degrading capacity; however, its application is hindered by the lack of fundamental gene expression tools. Here, we present a synthetic biology toolkit that enables rapid creation of functional gene circuits in C. testosteroni. We first built a shuttle system that allows efficient circuit construction in E. coli and necessary phenotypic testing in C. testosteroni. Then, we tested a set of wildtype inducible promoters, and further used a hybrid strategy to create engineered promoters to expand expression strength and dynamics. Additionally, we tested the T7 RNA Polymerase-PT7 promoter system and reduced its leaky expression through promoter mutation for gene expression. By coupling random library construction with FACS screening, we further developed a synthetic T7 promoter library to confer a wider range of expression strength and dynamic characteristics. This study provides a set of valuable tools to engineer gene circuits in C. testosteroni, facilitating the establishment of the organism as a useful microbial chassis for bioremediation purposes.
KW - Comamonas testosteroni
KW - P promoter library
KW - T7 RNA polymerase-P system
KW - inducible promoter
KW - shuttle vehicle
KW - synthetic biology toolkit
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UR - http://www.scopus.com/inward/citedby.url?scp=85048128443&partnerID=8YFLogxK
U2 - 10.1021/acssynbio.7b00430
DO - 10.1021/acssynbio.7b00430
M3 - Article
VL - 7
SP - 1753
EP - 1762
JO - ACS Synthetic Biology
JF - ACS Synthetic Biology
SN - 2161-5063
IS - 7
ER -