In vitro elaboration Mutagenesis and cloning of the PA gene in Bacillus subtilis

Abstract

Background: The immune antigen of Bacillus anthracis is a protein that can attach to the surface receptor of all human cells. At the surface of cancer cells, there is a receptor that activates the uPA (Urokinase plasminogen) that do not exist in normal human cells. Objectives: The aim of this study was changing the location of the attachment of the PA gene by a directed mutation in order to attach only to the cancer cells.Methods: PA gene was extracted from the pMNA1 plasmid. The mutation on the PA gene was made by Overlap Extension PCR. The mutated segment was transferred to DH5α; the strain of Escherichia coli. With TA coning carrier. By restriction enzymes Hind III and BamH I the mutated PA gene was extracted and transferred to pWB980 and by electroporation method, it was transferred to the WB600 strain.Results: In this study, the mutation was occurred in sequences of PA gene by SOE PCR method resulting in a change in the genetic code of amino acid 194. The occurrence of mutation was confirmed by determining base sequences. Conclusion: Cancer is a severe disease that has a major impact on large groups of people which the problem of cancer is a leading cause of death across the world. One of the treatment methods of cancer is bacterial toxins if only cancer cells receive them. Therefore, these mutated PA proteins can be effective as novel therapeutic agents for the treatment of cancer.