BACKGROUND: Salmonellosis is a dangerous disease that can threaten the health of humans and animals. This disease can lead to economic losses annually; therefore, many studies have been conducted to prevent this disease.OBJECTIVES: The current study aims to design a recombinant chimera protein HBHA-Omp28 as a vaccine against Salmonella typhimurium.METHODS: The nucleotide and amino acid sequences of Omp28 and HBHA proteins were first extracted from the NCBI database. Then, the recombinant chimera of HBHA-Omp28 was bioinformatically assembled using a rigid linker. Epitope prediction of T and B cells, antigenicity, allergenicity, and physicochemical features assessments of HBHA-Omp28 were done using Immune Epitope Database (IEDB), ABCpred, VaxiJen, AllerTOP and ProtParam online servers, respectively. To assess the secondary and tertiary structures, the Self-Optimized Prediction Method with Alignment (SOPMA) and the Iterative Threading ASSEmbly Refinement (I-TASSER) server were used, respectively. Molecular docking between recombinant chimera and TLR4/MD2 receptor was assessed by ClusPro server. Finally, after codon optimization of nucleotide sequence of recombinant chimera to express in Escherichia Coli k-12 strain, the cloning of recombinant chimera in pET21-a (+) vector was examined.RESULTS: The designed recombinant chimera was classified as an antigenic and non-allergenic protein with molecular weight of 34.19 kDa. According to the results of molecular docking study, the HBHA-Omp28 protein was able to bind to TLR4/MD2 receptor using 9 hydrogen bonds. The results of cloning study demonstrated that HBHA-Omp28 successfully cloned into pET21-a (+).CONCLUSIONS: The designed recombinant chimera can be an appropriate vaccine against salmonella bacteria.