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Host-Vector Systems for Amino Acid-Producing Coryneform Bacteria
Published in Yoshikatsu Murooka, Tadayuki Imanaka, Recombinant Microbes for Industrial and Agricultural Applications, 2020
To clone the genes encoding enzymes for amino acid biosynthesis, auxotrophic mutants are important for selection. In addition, as a general host-vector system, the reduction of restriction activity on foreign DNA is also very important to obtain a high transformation frequency. Thus, we tried to develop a restriction-deficient mutant by NTG mutagenesis. As the model of the foreign gene, we used virulent phage F-1A, which can propagate inB. flavum as well as inB. lactofermentum. The strategy was as follows: the restriction-deficient B. lactofermentum mutant, induced by NTG treatment, could be selected as that which allowed the propagation of the B. flavum-àzûvtà phage F-1A, whereas the wild-type B. lactofermentum did not. The r-43 thus obtained was transformed at a frequency of 3.9 x 103/μg pAJ655 DNA propagated in E. coli, whereas the wild-type B. lactofermentum ATCC 13869 was transformed at only 4 x 101 [37].
Selection and Improvement of Industrial Organisms for Biotechnological Applications
Published in Nduka Okafor, Benedict C. Okeke, Modern Industrial Microbiology and Biotechnology, 2017
Nduka Okafor, Benedict C. Okeke
Auxotrophic mutants are those which lack the enzymes to manufacture certain required nutrients. Consequently, such nutrients must therefore be added to the growth medium. In contrast, the wild-type or prototrophic organisms possess all the enzymes needed to synthesize all growth requirements. Auxotrophic mutants are often used in industrial microbiology, e.g. for the production of amino acids, nucleotides, etc.
Glossary of scientific and technical terms in bioengineering and biological engineering
Published in Megh R. Goyal, Scientific and Technical Terms in Bioengineering and Biological Engineering, 2018
Auxotroph is a mutant cell or micro-organism lacking one metabolic pathway present in the parental strain, and that consequently will not multiply on a minimal medium, but requires for growth the addition of a specific compound, such as an amino acid or a vitamin.
Biosafety and biosecurity in Synthetic Biology: A review
Published in Critical Reviews in Environmental Science and Technology, 2019
Lucía Gómez-Tatay, José M. Hernández-Andreu
Biocontainment measures include physical measures (engineering design of equipment, process and production plant) and biological means, among which inducible systems, auxotrophy and cellular circuits are the most established strategies. In inducible systems, the introduced genes are expressed only if a specific inducer is present. The inducer required is not common in the natural environment, so that if the organism escapes the lab, the engineered trait will not be expressed, avoiding any potential advantage given by the genetic construct. Auxotrophy implies that the organism is not able to produce a particular vital compound, which must be provided in its media. Cellular circuits are genetic constructs that can lead to cell death when activated (kill switches) or to the death of the new host in case of horizontal gene transfer (addiction modules). However, these strategies are not effective enough when they are used in isolation, since the evolutionary cost of bypassing or reverting the containment mechanism is very low. Multiple strategies and targets must therefore be combined, in what is called multi-layered containment (Torres, Krüger, Csibra, Gianni, & Pinheiro, 2016), although it must be taken into account that multi-layering increases complexity, thus making the device more prone to failure. Furthermore, precautions must be adopted such as not to incorporate antibiotic-resistance genes as markers for plasmid selection, in order to avoid their propagation (Wright, Stan, & Ellis, 2013). To date, there have been no reports in the scientific literature of an escape of synthetic organisms into the natural environment.