Expression of Clostridium difficile toxin A receptor binding domain in Lactococcus lactis
AIM: To expreess the Clostridium difficile toxin A receptor binding domain and test its bioactivity. METHODS: The gene of fourteen of the 38 C-terminal toxin A repeats (14CDTA) coding for the clostridium difficile toxin A receptor binding domain was amplified and then cloned into the lactococcus lactis expression system pNBC1000 and pNBC2000 to form pNBC1001 and pNBC2001, the confirmed sequence was cut and ligated to pTRKH2 a shuttle vector for E. coli and Lactococcus lactis to generate pNBCL1001 and pNBCL2001. pNBCL1001 and pNBCL2001 were transformed into Lactococcus lactis by electroporation. Protein electrophoresis and Western-Blot were used to detect the expession of 14CDTA and its bioactivity. RESULTS: The 14CDTA expression vectors were constructed successfully. The secretory form of 14CDTA had a weight of 39.2 ku and the cell anchored form of 14CDTA had a weight of 55.1 ku. All the forms of 14CDTA could be deteced with the clostridium difficile toxin A multicloned antibody by Western Blotting. CONCLUSION: The recombinant lactococcus lactis expressing the 14CDTA may lay a foundation for vaccine against Clostridium difficile.
NETL
NSTL
万方数据
维普
