SUMMARYPseudomonas arvillamt-2 (ATCC 23073) has been shown to harbour a transmissible plasmid which codes for the degradation of benzoate andm-toluate. Plasmid-borne genetic information codes for the conversion of these compounds to catechol then the assimilation of catechol via themetacleavage pathway.
SUMMARYThe TOL(M1) metabolic plasmid was transferred fromPseudomonas arvillamt-2 to a mutant ofPseudomonas putida. Although neither the donor nor the recipient could grow on phenol, the transconjugants could grow slowly on this carbon source. In these transconjugants phenol was converted to catechol by chromosomal encoded phenol hydroxylase followed by degradation of catechol by low uninduced levels of the plasmid encoded catecholmetacleavage pathway. A mutant, which grew well on phenol, was isolated from one of the transconjugants and it was found that phenol could now act as an inducer for themetacleavage pathway.
The ability to efficiently and reliably transfer genetic circuits between the key synthetic biology chassis, such as Escherichia coli and Bacillus subtilis, constitutes one of the major hurdles of the rational genome engineering. Using lambda Red recombineering we integrated the thermosensitive lambda repressor and the lysis genes of several bacteriophages into the E. coli chromosome. The lysis of the engineered autolytic cells is inducible by a simple temperature shift. We improved the lysis efficiency by introducing different combinations of lysis genes from bacteriophages lambda, ΦX174 and MS2 under the control of the thermosensitive lambda repressor into the E. coli chromosome. We tested the engineered autolytic cells by transferring plasmid and bacterial artificial chromosome (BAC)-borne genetic circuits from E. coli to B. subtilis. Our engineered system combines benefits of the two main synthetic biology chassis, E. coli and B. subtilis, and allows reliable and efficient transfer of DNA edited in E. coli into B. subtilis.
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