A genetic toolkit and gene switches to limit Mycoplasma growth for a synthetic vaccine chassis

2021 ◽  
Author(s):  
Alicia Broto ◽  
Erika Gaspari ◽  
Samuel Miravet-Verde ◽  
Vitor Martins dos Santos ◽  
Mark Isalan
Keyword(s):  
Synthetic Gene ◽  
Minimal Cell ◽  
Essential Nutrients ◽  
Gene Circuits ◽  
Live Attenuated Vaccines ◽  
Synthetic Gene Circuits ◽  
Gene Switches ◽  
Genetic Toolkit

Abstract Mycoplasmas have exceptionally streamlined genomes and are strongly adapted to their many hosts, which provide them with essential nutrients. Owing to their relative genomic simplicity, Mycoplasmas have been used for the development of chassis to deploy tailored vaccines. However, the dearth of robust and precise toolkits for genomic manipulation and tight regulation has hindered any substantial advance. Herein we describe the construction of a robust genetic toolkit for M. pneumoniae, and its successful deployment to engineer synthetic gene switches that control and limit Mycoplasma growth, for biosafety containment applications. We found these synthetic gene circuits to be stable and robust in the long-term, in the context of a minimal cell. With this work, we lay a foundation to develop viable and robust biosafety systems to exploit a synthetic Mycoplasma chassis for live attenuated vaccines or even for live vectors for biotherapeutics.

Synthetic Gene Circuits: Design, Implement, and Apply

2021 ◽  
pp. 1-18
Author(s):  
Andrew Lezia ◽  
Arianna Miano ◽  
Jeff Hasty
Keyword(s):  
Synthetic Gene ◽  
Gene Circuits ◽  
Synthetic Gene Circuits

Synthetic Gene Circuits

2014 ◽  
pp. 1-56 ◽  
Author(s):  
Barbara Jusiak ◽  
Ramiz Daniel ◽  
Fahim Farzadfard ◽  
Lior Nissim ◽  
Oliver Purcell ◽  
...  
Keyword(s):  
Synthetic Gene ◽  
Gene Circuits ◽  
Synthetic Gene Circuits

scholarly journals Two cellular resource‐based models linking growth and parts characteristics aids the study and optimisation of synthetic gene circuits

2017 ◽  
Vol 1 (1) ◽  
pp. 30-39 ◽  
Author(s):  
Huijuan Wang ◽  
Maurice H.T. Ling ◽  
Tze Kwang Chua ◽  
Chueh Loo Poh
Keyword(s):  
Synthetic Gene ◽  
Gene Circuits ◽  
Synthetic Gene Circuits

scholarly journals Multistable and dynamic CRISPRi-based synthetic circuits

2019 ◽  
Author(s):  
Javier Santos-Moreno ◽  
Eve Tasiudi ◽  
Joerg Stelling ◽  
Yolanda Schaerli
Keyword(s):  
Context Dependency ◽  
Synthetic Gene ◽  
Toggle Switch ◽  
Stripe Pattern ◽  
Gene Circuits ◽  
Synthetic Gene Circuits ◽  
Gene Expression Control ◽  
Unspecific Binding ◽  
High Predictability ◽  
Pattern Forming

AbstractGene expression control based on CRISPRi (clustered regularly interspaced short palindromic repeats interference) has emerged as a powerful tool for creating synthetic gene circuits, both in prokaryotes and in eukaryotes; yet, its lack of cooperativity has been pointed out as a potential obstacle for dynamic or multistable circuit construction. Here we use CRISPRi to build prominent synthetic gene circuits in Escherichia coli. We report the first-ever CRISPRi oscillator (“CRISPRlator”), bistable network (toggle switch) and stripe pattern-forming incoherent feed-forward loop (IFFL). Our circuit designs, conceived to feature high predictability and orthogonality, as well as low metabolic burden and context-dependency, allowed us to achieve robust circuit behaviors. Mathematical modeling suggests that unspecific binding in CRISPRi is essential to establish multistability. Our work demonstrates the wide applicability of CRISPRi in synthetic circuits and paves the way for future efforts towards engineering more complex synthetic networks, boosted by the advantages of CRISPR technology.

scholarly journals Synthetic Gene Circuits for Antimicrobial Resistance and Cancer Research

2021 ◽  
Author(s):  
Kevin S. Farquhar ◽  
Michael Tyler Guinn ◽  
Gábor Balázsi ◽  
Daniel A. Charlebois
Keyword(s):  
Mammalian Cells ◽  
Rational Design ◽  
Study Drug ◽  
Quantitative Model ◽  
Synthetic Gene ◽  
Modeling Tools ◽  
Gene Circuits ◽  
Novel Treatments ◽  
Synthetic Gene Circuits ◽  
Temporal Dimensions

Mathematical models and synthetic gene circuits are powerful tools to develop novel treatments for patients with drug-resistant infections and cancers. Mathematical modeling guides the rational design of synthetic gene circuits. These systems are then assembled into unified constructs from existing and/or modified genetic components from a range of organisms. In this chapter, we describe modeling tools for the design and characterization of chemical- and light-inducible synthetic gene circuits in different organisms and highlight how synthetic gene circuits are advancing biomedical research. Specifically, we demonstrate how these quantitative model systems are being used to study drug resistance in microbes and to probe the spatial–temporal dimensions of cancer in mammalian cells.

scholarly journals Characterizing microRNA-mediated modulation of gene expression noise and its effect on synthetic gene circuits

2020 ◽  
Author(s):  
Lei Wei ◽  
Shuailin Li ◽  
Tao Hu ◽  
Michael Q. Zhang ◽  
Zhen Xie ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Types ◽  
Classification Performance ◽  
Synthetic Gene ◽  
Gene Circuits ◽  
Gene Expression Noise ◽  
Expression Noise ◽  
Synthetic Gene Circuits ◽  
Highly Expressed Genes ◽  
Lower Noise

AbstractGene expression noise plays an important role in many biological processes, such as cell differentiation and reprogramming. It can also dramatically influence the behavior of synthetic gene circuits. MicroRNAs (miRNAs) have been shown to reduce the noise of lowly expressed genes and increase the noise of highly expressed genes, but less is known about how miRNAs with different properties may regulate gene expression noise differently. Here, by quantifying gene expression noise using mathematical modeling and experimental measurements, we showed that competing RNAs and the composition of miRNA response elements (MREs) play important roles in modulating gene expression noise. We found that genes targeted by miRNAs with weak competing RNAs show lower noise than those targeted by miRNAs with strong competing RNAs. In addition, in comparison with a single MRE, repetitive MREs targeted by the same miRNA suppress the noise of lowly expressed genes but increase the noise of highly expressed genes. Additionally, MREs composed of different miRNA targets could cause similar repression levels but lower noise compared with repetitive MREs. We further observed the influence of miRNA-mediated noise modulation in synthetic gene circuits which could be applied to classify cell types using miRNAs as sensors. We found that miRNA sensors that introduce higher noise could lead to better classification performance. Our results provide a systematic and quantitative understanding of the function of miRNAs in controlling gene expression noise and how we can utilize miRNAs to modulate the behavior of synthetic gene circuits.

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