An effective method to develop a safe vaccine against botulism is to utilize molecular biology techniques to produce recombinant antigens, which provoke the immune response in the recipient organism. A suggested antigen is a specific recombinant fragment of the botulinum neurotoxin (BoNT), which elicits the predictable immune response and does not have any toxic effects. In this study, the binding domain of the heavy chain of BoNT serotype A, which is the responsible subunit for binding to the

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... eceptor(s) of presynaptic membranes in neuromuscular junctions, is the selected fragment of this toxin to be recombinantly produced. Objectives: In order to prevent a severe syndrome such as Botulism, developing efficient vaccines against it is a necessity. Efforts have been made to accomplish this throughout time; however, some have discontinued due to the risks and unreliability of their production and usage. Methods: The encoding gene of BoNT/A-Hc was cloned into two different strains of Pichia pastoris, which were compared to each other based on the yield of the recombinant product. Results: The results demonstrated that the expression of recombinant BoNT/A-Hc by PichiaPink strain was successful, and the achieved recombinant BoNT/A-Hc was subsequently purified and then verified by using the specific antibody and analytical methods. Conclusions: In contrast, the expression results from the X-33 strain were not significant.