Enabling plant synthetic biology through genome engineering

2015 ◽  
Vol 33 (2) ◽  
pp. 120-131 ◽  
Author(s):  
Nicholas J. Baltes ◽  
Daniel F. Voytas
Keyword(s):  
Synthetic Biology ◽  
Genome Engineering ◽  
Plant Synthetic Biology

Genome Engineering Tools in Plant Synthetic Biology

2019 ◽  
pp. 47-73 ◽  
Author(s):  
Krishan Mohan Rai ◽  
Kaushik Ghose ◽  
Anshulika Rai ◽  
Harpal Singh ◽  
Rakesh Srivastava ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Genome Engineering ◽  
Plant Synthetic Biology

GondenBraid2.0: A comprehensive toolkit for plant synthetic biology

Planta Medica ◽  
2013 ◽  
Vol 79 (13) ◽  
Author(s):  
A Sarrion-Perdigones ◽  
M Vazquez-Vilar ◽  
J Palaci ◽  
A Granell ◽  
D Orzáez
Keyword(s):  
Synthetic Biology ◽  
Plant Synthetic Biology

scholarly journals Development of oriC-Based Plasmids for Mesoplasma florum

2017 ◽  
Vol 83 (7) ◽  
Author(s):  
Dominick Matteau ◽  
Marie-Eve Pepin ◽  
Vincent Baby ◽  
Samuel Gauthier ◽  
Mélissa Arango Giraldo ◽  
...  
Keyword(s):  
Systems Biology ◽  
Synthetic Biology ◽  
Genetic Engineering ◽  
Genome Engineering ◽  
Antibiotic Resistance Genes ◽  
Viable Cell ◽  
Pathogenic Potential ◽  
Strong Basis ◽  
Content Type ◽  
Basic Biology

ABSTRACT The near-minimal bacterium Mesoplasma florum constitutes an attractive model for systems biology and for the development of a simplified cell chassis in synthetic biology. However, the lack of genetic engineering tools for this microorganism has limited our capacity to understand its basic biology and modify its genome. To address this issue, we have evaluated the susceptibility of M. florum to common antibiotics and developed the first generation of artificial plasmids able to replicate in this bacterium. Selected regions of the predicted M. florum chromosomal origin of replication (oriC) were used to create different plasmid versions that were tested for their transformation frequency and stability. Using polyethylene glycol-mediated transformation, we observed that plasmids harboring both rpmH-dnaA and dnaA-dnaN intergenic regions, interspaced or not with a copy of the dnaA gene, resulted in a frequency of ∼4.1 × 10−6 transformants per viable cell and were stably maintained throughout multiple generations. In contrast, plasmids containing only one M. florum oriC intergenic region or the heterologous oriC region of Mycoplasma capricolum, Mycoplasma mycoides, or Spiroplasma citri failed to produce any detectable transformants. We also developed alternative transformation procedures based on electroporation and conjugation from Escherichia coli, reaching frequencies up to 7.87 × 10−6 and 8.44 × 10−7 transformants per viable cell, respectively. Finally, we demonstrated the functionality of antibiotic resistance genes active against tetracycline, puromycin, and spectinomycin/streptomycin in M. florum. Taken together, these valuable genetic tools will facilitate efforts toward building an M. florum-based near-minimal cellular chassis for synthetic biology. IMPORTANCE Mesoplasma florum constitutes an attractive model for systems biology and for the development of a simplified cell chassis in synthetic biology. M. florum is closely related to the mycoides cluster of mycoplasmas, which has become a model for whole-genome cloning, genome transplantation, and genome minimization. However, M. florum shows higher growth rates than other Mollicutes, has no known pathogenic potential, and possesses a significantly smaller genome that positions this species among some of the simplest free-living organisms. So far, the lack of genetic engineering tools has limited our capacity to understand the basic biology of M. florum in order to modify its genome. To address this issue, we have evaluated the susceptibility of M. florum to common antibiotics and developed the first artificial plasmids and transformation methods for this bacterium. This represents a strong basis for ongoing genome engineering efforts using this near-minimal microorganism.

scholarly journals The Multiplanetary Future of Plant Synthetic Biology

Genes ◽  
2018 ◽  
Vol 9 (7) ◽  
pp. 348 ◽  
Author(s):  
Briardo Llorente ◽  
Thomas Williams ◽  
Hugh Goold
Keyword(s):  
Synthetic Biology ◽  
Plant Synthetic Biology

scholarly journals Streamlined Construction of the Cyanobacterial CO2-Fixing Organelle via Protein Domain Fusions for Use in Plant Synthetic Biology

2015 ◽  
Vol 27 (9) ◽  
pp. 2637-2644 ◽  
Author(s):  
C. Raul Gonzalez-Esquer ◽  
Tyler B. Shubitowski ◽  
Cheryl A. Kerfeld
Keyword(s):  
Synthetic Biology ◽  
Protein Domain ◽  
Plant Synthetic Biology

scholarly journals Attenuation of a Pathogenic Mycoplasma Strain by Modification of the obg Gene by Using Synthetic Biology Approaches

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Carole Lartigue ◽  
Yanina Valverde Timana ◽  
Fabien Labroussaa ◽  
Elise Schieck ◽  
Anne Liljander ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Genome Engineering ◽  
Recipient Cell ◽  
Single Point ◽  
Chemical Mutagenesis ◽  
Economic Losses ◽  
Temperature Sensitive ◽  
Content Type ◽  
Mycoplasma Mycoides ◽  
Virulent Phenotype

ABSTRACT Mycoplasma species are responsible for several economically significant livestock diseases for which there is a need for new and improved vaccines. Most of the existing mycoplasma vaccines are attenuated strains that have been empirically obtained by serial passages or by chemical mutagenesis. The recent development of synthetic biology approaches has opened the way for the engineering of live mycoplasma vaccines. Using these tools, the essential GTPase-encoding gene obg was modified directly on the Mycoplasma mycoides subsp. capri genome cloned in yeast, reproducing mutations suspected to induce a temperature-sensitive (TS+) phenotype. After transplantation of modified genomes into a recipient cell, the phenotype of the resulting M. mycoides subsp. capri mutants was characterized. Single-point obg mutations did not result in a strong TS+ phenotype in M. mycoides subsp. capri, but a clone presenting three obg mutations was shown to grow with difficulty at temperatures of ≥40°C. This particular mutant was then tested in a caprine septicemia model of M. mycoides subsp. capri infection. Five out of eight goats infected with the parental strain had to be euthanized, in contrast to one out of eight goats infected with the obg mutant, demonstrating an attenuation of virulence in the mutant. Moreover, the strain isolated from the euthanized animal in the group infected with the obg mutant was shown to carry a reversion in the obg gene associated with the loss of the TS+ phenotype. This study demonstrates the feasibility of building attenuated strains of mycoplasma that could contribute to the design of novel vaccines with improved safety. IMPORTANCE Animal diseases due to mycoplasmas are a major cause of morbidity and mortality associated with economic losses for farmers all over the world. Currently used mycoplasma vaccines exhibit several drawbacks, including low efficacy, short time of protection, adverse reactions, and difficulty in differentiating infected from vaccinated animals. Therefore, there is a need for improved vaccines to control animal mycoplasmoses. Here, we used genome engineering tools derived from synthetic biology approaches to produce targeted mutations in the essential GTPase-encoding obg gene of Mycoplasma mycoides subsp. capri. Some of the resulting mutants exhibited a marked temperature-sensitive phenotype. The virulence of one of the obg mutants was evaluated in a caprine septicemia model and found to be strongly reduced. Although the obg mutant reverted to a virulent phenotype in one infected animal, we believe that these results contribute to a strategy that should help in building new vaccines against animal mycoplasmoses.

scholarly journals Synthetic Virus-Derived Nanosystems (SVNs) for Delivery and Precision Docking of Large Multifunctional DNA Circuitry in Mammalian Cells

Pharmaceutics ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 759
Author(s):  
Francesco Aulicino ◽  
Julien Capin ◽  
Imre Berger
Keyword(s):  
Gene Therapy ◽  
Synthetic Biology ◽  
Base Pair ◽  
Mammalian Cells ◽  
Viral Vectors ◽  
Genome Engineering ◽  
Dna Delivery ◽  
Attractive Alternative ◽  
Exogenous Dna ◽  
Nonviral Delivery

DNA delivery is at the forefront of current research efforts in gene therapy and synthetic biology. Viral vectors have traditionally dominated the field; however, nonviral delivery systems are increasingly gaining traction. Baculoviruses are arthropod-specific viruses that can be easily engineered and repurposed to accommodate and deliver large sequences of exogenous DNA into mammalian cells, tissues, or ultimately organisms. These synthetic virus-derived nanosystems (SVNs) are safe, readily customized, and can be manufactured at scale. By implementing clustered regularly interspaced palindromic repeats (CRISPR) associated protein (CRISPR/Cas) modalities into this system, we developed SVNs capable of inserting complex DNAs into genomes, at base pair precision. We anticipate a major role for SVNs as an attractive alternative to viral vectors in accelerating genome engineering and gene therapy applications in the future.

scholarly journals Standards for plant synthetic biology: a common syntax for exchange ofDNAparts

2015 ◽  
Vol 208 (1) ◽  
pp. 13-19 ◽  
Author(s):  
Nicola J. Patron ◽  
Diego Orzaez ◽  
Sylvestre Marillonnet ◽  
Heribert Warzecha ◽  
Colette Matthewman ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Plant Synthetic Biology

scholarly journals Base editors: modular tools for the introduction of point mutations in living cells

2019 ◽  
Vol 3 (5) ◽  
pp. 483-491 ◽  
Author(s):  
Mallory Evanoff ◽  
Alexis C. Komor
Keyword(s):  
Synthetic Biology ◽  
Genome Editing ◽  
Genome Engineering ◽  
Point Mutations ◽  
Living Cells ◽  
Single Base ◽  
New Family ◽  
Alternative Techniques ◽  
Cas Proteins ◽  
Modular Nature

Base editors are a new family of programmable genome editing tools that fuse ssDNA (single-stranded DNA) modifying enzymes to catalytically inactive CRISPR-associated (Cas) endonucleases to induce highly efficient single base changes. With dozens of base editors now reported, it is apparent that these tools are highly modular; many combinations of ssDNA modifying enzymes and Cas proteins have resulted in a variety of base editors, each with its own unique properties and potential uses. In this perspective, we describe currently available base editors, highlighting their modular nature and describing the various options available for each component. Furthermore, we briefly discuss applications in synthetic biology and genome engineering where base editors have presented unique advantages over alternative techniques.

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