Construction of Genetic Logic Gates Based on the T7 RNA Polymerase Expression System in Rhodococcus opacus PD630

ACS Synthetic Biology
Drew M DeLorenzo, Tae Seok Moon

Abstract

Rhodococcus opacus PD630 (R. opacus) is a nonmodel, Gram-positive bacterium that holds promise as a biological catalyst for the conversion of lignocellulosic biomass to value-added products. In particular, it demonstrates both a high tolerance for and an ability to consume inhibitory lignin-derived aromatics, generates large quantities of lipids, exhibits a relatively rapid growth rate, and has a growing genetic toolbox for engineering. However, the availability of genetic parts for tunable, high-activity gene expression is still limited in R. opacus. Furthermore, genetic logic circuits for sophisticated gene regulation have never been demonstrated in Rhodococcus spp. To address these shortcomings, two inducible T7 RNA polymerase-based expression systems were implemented for the first time in R. opacus and applied to the construction of AND and NAND genetic logic gates. Additionally, three isopropyl β-d-1-thiogalactopyranoside (IPTG)-inducible promoters were created by inserting LacI binding sites into newly characterized constitutive promoters. Furthermore, four novel aromatic sensors for 4-hydroxybenzoic acid, vanillic acid, sodium benzoate, and guaiacol were developed, expanding the gene expression toolbox. Finally, the T7 R...Continue Reading

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Citations

Dec 21, 2019·Biotechnology Journal·Austin G RottinghausTae Seok Moon
Oct 23, 2019·Biotechnology for Biofuels·Guadalupe Alvarez-Gonzalez, Neil Dixon
May 11, 2020·Current Opinion in Chemical Biology·Kirsten Davis, Tae Seok Moon
Apr 1, 2021·ACS Synthetic Biology·Drew M DeLorenzoTae Seok Moon
Apr 7, 2021·Biotechnology Advances·Youxiang Liang, Huimin Yu
Jul 5, 2021·Journal of Hazardous Materials·Kha Mong TranDokyun Na
Aug 26, 2021·Synthetic and Systems Biotechnology·Youxiang LiangHuimin Yu

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