Abstract: Highly conserved spvB gene is an important virulence factor of infected cells apoptosis, which encoded product with the ADPribosyltransferaseenzymatic activity. To further study the pathogenic mechanism of the spvB gene, this study began to build a mutant of the spvB gene in Salmonella typhimurium. According to the spvB gene sequence in GeneBank, two pair’s primers were designed with Primer Premier 5.0, upper- and down-stream of the spvB gene to amplify two homologous DNA fragments. The homologous recombinant DNA fragment of the defective target gene was cloned into the suicide plasmid PGMB151, which was then transduced into the target cell of S. typhimurium SR-11 for homologous recombination. The recombination was identified by PCR. A deletion mutant of the spvB gene in SR-11 was confirmed by PCR and sequencing analysis.