Toxin-antitoxin system

Keep your plasmids without antibiotic resistance

One of the challenges when creating synthetic systems in bacteria that serve a purpose besides increasing the fitness of the organism is that there is a negative selective pressure against keeping the system. Toxin-antitoxin systems can be used to make plasmids far more stabile without having to use antibiotics and antibiotic resistance. If a clone were to lose the plasmid, the toxin which usually has a longer half life than the antitoxin will kill the bacteria.

Above is an example of how a toxin-antitoxin system could be applied together with a gene X. If the gene is toxic or expressed strongly enough there will be a substantial evolutionary pressure to lose the plasmid during cell division. However if the gene is present on a plasmid with a toxin-antitoxin system would be lethal due to the loss of the antitoxin gene.

A natural toxin-antitoxin from lactobacillus plantarum

We have taken the Pem toxin-antitoxin system from plasmid p256 that was originally isolated from lactobacillus plantarum NC7. The system consists of a single operon and consists of two ORFs, one for the toxin and antitoxin respectively. Pem on p256 has been shown to increase segregational stability under non-selective pressure. The system has experimentally been shown to allow 88-100% retention of a plasmid after 80 generations(1). We have provided the toxin-antitoxin system both with and without a natural putative promoter.

References:

http://mic.sgmjournals.org/content/151/2/421.long (1)