Recent advances in plasmid-based tools for establishing novel microbial chassis

Nora, Luísa Czamanski; Westmann, Cauã Antunes; Guazzaroni, María Eugenia; Siddaiah, Chandranayaka; Gupta, Vijai Kumar; Silva‐Rocha, Rafael

doi:10.1016/j.biotechadv.2019.107433

Cited by 26 publications

(24 citation statements)

References 297 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Based on our experience, the cost of creation of these libraries falls in the range of $4,000-$12,000 per library, and it takes about 3 mo on average for building fully characterized libraries. The newly developed sequencing technologies and extendibility of genetic tools to nonmodel organisms can further reduce the time and cost of generating the LOF and GOF libraries [70,210,211]. Once the libraries are built, the running cost of these genome-wide assays across 96 conditions cost about $10 per assay and serve as a standardized reagent tool for in-house experiments or for sharing and comparing data across different laboratories.…”

Section: Discussionmentioning

confidence: 99%

High-throughput mapping of the phage resistance landscape in E. coli

et al. 2020

View full text Add to dashboard Cite

Bacteriophages (phages) are critical players in the dynamics and function of microbial communities and drive processes as diverse as global biogeochemical cycles and human health. Phages tend to be predators finely tuned to attack specific hosts, even down to the strain level, which in turn defend themselves using an array of mechanisms. However, to date, efforts to rapidly and comprehensively identify bacterial host factors important in phage infection and resistance have yet to be fully realized. Here, we globally map the host genetic determinants involved in resistance to 14 phylogenetically diverse double-stranded DNA phages using two model Escherichia coli strains (K-12 and BL21) with known sequence divergence to demonstrate strain-specific differences. Using genome-wide lossof-function and gain-of-function genetic technologies, we are able to confirm previously described phage receptors as well as uncover a number of previously unknown host factors that confer resistance to one or more of these phages. We uncover differences in resistance factors that strongly align with the susceptibility of K-12 and BL21 to specific phage. We also identify both phage-specific mechanisms, such as the unexpected role of cyclic-di-GMP in host sensitivity to phage N4, and more generic defenses, such as the overproduction of colanic acid capsular polysaccharide that defends against a wide array of phages. Our results indicate that host responses to phages can occur via diverse cellular mechanisms. Our systematic and high-throughput genetic workflow to characterize phage-host interaction determinants can be extended to diverse bacteria to generate datasets that allow predictive models of how phage-mediated selection will shape bacterial phenotype and evolution. The results of this study and future efforts to map the phage resistance landscape will lead to new insights into the coevolution of hosts and their phage, which can

show abstract

Section: Discussionmentioning

confidence: 99%

High-throughput mapping of the phage resistance landscape in E. coli

et al. 2020

View full text Add to dashboard Cite

show abstract

“…The genetic tools used in this work adhere to the genetic BioBrick standard, thereby allowing to rapidly generate, fuse and exchange biological parts to fulfill certain functions [61]. Furthermore, applying standardized genetic parts allows their use in a broad variety of different chassis, beyond the use of B. subtilis, and opens up the creation of biohybrids with novel functionalities [62]. The modularity of this concept is of particular importance, since it allows to rapidly implement the engineering driven principles such as abstraction and standardization [63].…”

Section: Discussionmentioning

confidence: 99%

Genetically Engineered Bacterial Biohybrid Microswimmers for Sensing Applications

Sun

Loderer

Revilla-Guarinos

2019

Sensors

View full text Add to dashboard Cite

Bacterial biohybrid microswimmers aim at exploiting the inherent motion capabilities of bacteria (carriers) to transport objects (cargoes) at the microscale. One of the most desired properties of microswimmers is their ability to communicate with their immediate environment by processing the information and producing a useful response. Indeed, bacteria are naturally equipped with such communication skills. Hereby, two-component systems (TCSs) represent the key signal transducing machinery and enable bacteria to sense and respond to a variety of stimuli. We engineered a natural microswimmer based on the Gram-positive model bacterium Bacillus subtilis for the development of biohybrids with sensing abilities. B. subtilis naturally adhered to silica particles, giving rise to different motile biohybrids systems with variable ratios of carrier(s)-to-cargo(es). Genetically engineered TCS pathways allowed us to couple the binding to the inert particles with signaling the presence of antibiotics in their surroundings. Activation of the antibiotic-induced TCSs resulted in fluorescent bacterial carriers as a response readout. We demonstrate that the genetically engineered TCS-mediated signaling capabilities of B. subtilis allow for the custom design of bacterial hybrid microswimmers able to sense and signal the presence of target molecules in the environment. The generally recognized as safe (GRAS) status of B. subtilis makes it a promising candidate for human-related applications of these novel biohybrids.

show abstract

“…This recovery of function, however, requires that photosynthesislethal microalgal mutants, which act as the transgene recipient strain, can be propagated heterotrophically. Alternatively, a recombinase has been applied to remove marker genes in engineered cyanobacteria [191]. Extrachromosomal vectors, or episomes, offer another tool for "non-GM" breeding in microalgae [192].…”

Section: From Nucleus To Organelle Engineeringmentioning

confidence: 99%

Engineering microalgae: transition from empirical design to programmable cells

Zhang

et al. 2021

Critical Reviews in Biotechnology

View full text Add to dashboard Cite

Domesticated microalgae hold great promise for the sustainable provision of various bioresources for human domestic and industrial consumption. Efforts to exploit their potential are far from being fully realized due to limitations in the know-how of microalgal engineering. The associated technologies are not as well developed as those for heterotrophic microbes, cyanobacteria, and plants. However, recent studies on microalgal metabolic engineering, genome editing, and synthetic biology have immensely helped to enhance transformation efficiencies and are bringing new insights into this field. Therefore, this article, summarizes recent developments in microalgal biotechnology and examines the prospects for generating specialty and commodity products through the processes of metabolic engineering and synthetic biology. After a brief examination of empirical engineering methods and vector design, this article focuses on quantitative transformation cassette design, elaborates on target editing methods and emerging digital design of algal cellular metabolism to arrive at high yields of valuable products. These advances have enabled a transition of manners in microalgal engineering from single-gene and enzyme-based metabolic engineering to systems-level precision engineering, from cells created with genetically modified (GM) tags to that without GM tags, and ultimately from proof of concept to tangible industrial applications. Finally, future trends are proposed in microalgal engineering, aiming to establish individualized transformation systems in newly identified species for strain-specific specialty and commodity products, while developing sophisticated universal toolkits in model algal species.

show abstract

Recent advances in plasmid-based tools for establishing novel microbial chassis

Cited by 26 publications

References 297 publications

High-throughput mapping of the phage resistance landscape in E. coli

High-throughput mapping of the phage resistance landscape in E. coli

Genetically Engineered Bacterial Biohybrid Microswimmers for Sensing Applications

Engineering microalgae: transition from empirical design to programmable cells

Contact Info

Product

Resources

About