The expression of biologically active cholera toxin in Escherichia coli

Chromosomal DNA from Vibrio cholerae El Tor strain 1621 was digested with Hind III and the products fractionated by centrifugatlon through a sucrose gradient. A 15kb fragment containing the toxin gene of V. cholerae was identified by its homology with the heat labile toxin (LT) gene of toxigenic E. coli. This fragment was cloned in E. coli using pATl53 and subsequently characterised by digestion with different restriction endonucleases. Sequences homologous to the LT gene were identified by hybridisation and then sub-cloned using either pATl53 or pACYC184. Expression of the cloned CT gene in E. coli was detected using both cell culture and ELISA assays. One recombinant plasmid coded for the synthesis of an immunologically active but biologically inactive derivative of CT.