Tetanus is a disease caused by tetanus toxin, a potent inhibitor for the release of inhibitory neurotransmitter in the central nervous system that causes spastic paralysis. Fragment C (52 kD) of this toxin is responsible for binding to the neuronal membrane. For this reason, and also its non toxigenic and immunogenic nature, this fragment might be ideal for new vaccine development. Presently, with respect to the incidence of disease in neonates and animals and the side effects of toxoid vaccine, designing a more effective and efficient vaccine for prevention of this disease is crucial. A segment of Clostridium tetani DNA corresponding to C fragment of tetanus toxin was amplified using polymerase chain reaction. This fragment was cloned into expression vector pMalc2x, under the control of the lac promoter. Expression of this plasmid in Escherichia coli was confirmed by western blotting. In this study, the vector had a strong promoter to allow high level expression of C fragment. Based on our results it appears that this recombinant plasmid may be suitable for the production and development of recombinant vaccine and also has many other applications, such as construction diagnostic kits, production hyperimmune antiserum for serotherapy and as a vehicle for drug delivery to CNS.