Purified cell-free systems as standard parts for synthetic biology

Matsubayashi, Hideaki; Ueda, Takuya

doi:10.1016/j.cbpa.2014.09.031

Cited by 35 publications

(27 citation statements)

References 30 publications

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“…Reconstitution of artificial living systems can shed light on the basic design rules of evolution and contribute to many new technologies such as in-vitro protein modification, engineering cellular behavior, as well as drug delivery, therapy and biosensing, all of which greatly promote the development of synthetic biology [ [131] , [132] , [133] ]. One of the most important applications of RNA replication in vitro is the study of rules that guide Darwinian evolution and building the smallest artificial living system ( Fig.…”

Section: Rna-replication-based Synthetic Systems Mimicking the Phage mentioning

confidence: 99%

Applications of phage-derived RNA-based technologies in synthetic biology

Zhang

2020

Synthetic and Systems Biotechnology

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As the most abundant biological entities with incredible diversity, bacteriophages (also known as phages) have been recognized as an important source of molecular machines for the development of genetic-engineering tools. At the same time, phages are crucial for establishing and improving basic theories of molecular biology. Studies on phages provide rich sources of essential elements for synthetic circuit design as well as powerful support for the improvement of directed evolution platforms. Therefore, phages play a vital role in the development of new technologies and central scientific concepts. After the RNA world hypothesis was proposed and developed, novel biological functions of RNA continue to be discovered. RNA and its related elements are widely used in many fields such as metabolic engineering and medical diagnosis, and their versatility led to a major role of RNA in synthetic biology. Further development of RNA-based technologies will advance synthetic biological tools as well as provide verification of the RNA world hypothesis. Most synthetic biology efforts are based on reconstructing existing biological systems, understanding fundamental biological processes, and developing new technologies. RNA-based technologies derived from phages will offer abundant sources for synthetic biological components. Moreover, phages as well as RNA have high impact on biological evolution, which is pivotal for understanding the origin of life, building artificial life-forms, and precisely reprogramming biological systems. This review discusses phage-derived RNA-based technologies terms of phage components, the phage lifecycle, and interactions between phages and bacteria. The significance of RNA-based technology derived from phages for synthetic biology and for understanding the earliest stages of biological evolution will be highlighted.

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Section: Rna-replication-based Synthetic Systems Mimicking the Phage mentioning

confidence: 99%

Applications of phage-derived RNA-based technologies in synthetic biology

Zhang

2020

Synthetic and Systems Biotechnology

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“…The cell-free protein expression system, especially the PURE system, has been recently applied to artificial cell models. The integration of de novo DNA synthesis with replication in the artificial cells provides genetic information sustainability in the synthetic system [ 111 ]. A critical step in artificial cell construction involves compartmentalized translation–transcription machinery materials or any biological elements from the exterior environment.…”

Section: Microfluidic Platforms and Cell-free Applicationsmentioning

confidence: 99%

Cell-Free Approaches in Synthetic Biology Utilizing Microfluidics

Damiati

Mhanna

Kodzius

et al. 2018

Genes

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Synthetic biology is a rapidly growing multidisciplinary branch of science which aims to mimic complex biological systems by creating similar forms. Constructing an artificial system requires optimization at the gene and protein levels to allow the formation of entire biological pathways. Advances in cell-free synthetic biology have helped in discovering new genes, proteins, and pathways bypassing the complexity of the complex pathway interactions in living cells. Furthermore, this method is cost- and time-effective with access to the cellular protein factory without the membrane boundaries. The freedom of design, full automation, and mimicking of in vivo systems reveal advantages of synthetic biology that can improve the molecular understanding of processes, relevant for life science applications. In parallel, in vitro approaches have enhanced our understanding of the living system. This review highlights the recent evolution of cell-free gene design, proteins, and cells integrated with microfluidic platforms as a promising technology, which has allowed for the transformation of the concept of bioprocesses. Although several challenges remain, the manipulation of biological synthetic machinery in microfluidic devices as suitable ‘homes’ for in vitro protein synthesis has been proposed as a pioneering approach for the development of new platforms, relevant in biomedical and diagnostic contexts towards even the sensing and monitoring of environmental issues.

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“…Recently, many researchers have proposed constructing artificial or minimal cell-like systems in vitro from biological molecules 1 – 5 . Reconstitution of cells and desired cellular functions from well-defined molecules enables us to understand the mechanisms or principles of target cellular functions 6 – 9 and to develop new technologies 10 – 13 .…”

Section: Introductionmentioning

confidence: 99%

Self-replication of circular DNA by a self-encoded DNA polymerase through rolling-circle replication and recombination

Sakatani

Yomo

Ichihashi

2018

Sci Rep

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A major challenge in constructing artificial cells is the establishment of a recursive genome replication system coupled with gene expression from the genome itself. One of the simplest schemes of recursive DNA replication is the rolling-circle replication of a circular DNA coupled with recombination. In this study, we attempted to develop a replication system based on this scheme using self-encoded phi29 DNA polymerase and externally supplied Cre recombinase. We first identified that DNA polymerization is significantly inhibited by Cre recombinase. To overcome this problem, we performed in vitro evolution and obtained an evolved circular DNA that can replicate efficiently in the presence of the recombinase. We also showed evidence that during replication of the evolved DNA, the circular DNA was reproduced through recombination by Cre recombinase. These results demonstrate that the evolved circular DNA can reproduce itself through gene expression of a self-encoded polymerase. This study provides a step forward in developing a simple recursive DNA replication system for use in an artificial cell.

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Purified cell-free systems as standard parts for synthetic biology

Cited by 35 publications

References 30 publications

Applications of phage-derived RNA-based technologies in synthetic biology

Applications of phage-derived RNA-based technologies in synthetic biology

Cell-Free Approaches in Synthetic Biology Utilizing Microfluidics

Self-replication of circular DNA by a self-encoded DNA polymerase through rolling-circle replication and recombination

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