CLONING AND EXPRESSION OF THE ZOT (ZONULA OCCLUDENS TOXIN) GENE OF Vibrio cholerae IN Escherichia coli

摘要

Two recombinant plasmids containing a cloned zot (zonula occludens toxin) gene of Vibrio cholerae, synthesized by the polymerase chain reaction, in an orientation ensuring a direction of transcription under the control of the lac promoter, were constructed. One of them also contained its own promoter of the zot gene. A fragment of the cholera vibrio DNA including intact zot and ace (accessory cholera enterotoxin) genes was also cloned in a third recombinant plasmid. When transformed into Escherichia coli, all three plasmids expressed the cloned genes with varying degrees of intensity, depending on the promoter, the host strain, and the conditions of cultivation. The possibility of using a CaCo2 intestinal epithelium cell culture to assess the biological action of the preparations of these toxins was shown. The product of the zot gene possess pronounced activity toward CaCo2, despite the absence of processing and transport of the toxin itself out of E. coli cells. The recombinant plasmids obtained can be used as a source of molecular probes, and E. coli clones that contain them can be used for isolation of Zot preparations for scientific and practical purposes.