In order to develop phage endolysin as a novel antibacterial agent for the prevention and control of Salmonella,the endolysin of Salmonella Typhimurium phage T139 was obtained by bioinformatics and heterologous expression of protein. The lytic activity of endolysin against Salmonella Typhimurium was verified. The genome of Salmonella Typhimurium phage T139 was analyzed and found to be 38 854 bp long,with an average GC content of 49.10% and no tRNA gene. The results of evolutionary tree analysis showed that phage T139 is belonged to the genus T7 phage with 43 ORFs predicted in total,23 ORFs being assigned known functions. The results of multiple sequence alignment and conserved structural domain analysis showed that the protein encoded by the orf 40 gene (LysT40) containing 126 amino acid residues with a conserved structural domain of phage endopeptidase,making it a possible phage endolysin. The presence of a large number of helical structures (70.63%) in the secondary structure of LysT40 allows the protein to maintain the stable structure. The endopeptidase gene orf 40 was heterologously expressed,purified and analyzed by SDS-PAGE and Western blot to obtain a single target protein band at 10-15 ku at a purified protein concentration of 280 μg/mL. LysT40 had significantly lytic activity against chloroform or EDTA pretreated Salmonella Typhimurium within 30 min,with absorbance values OD600 decreasing by 0.18 and 0.31 compared to the control. It will provide a basis for developing phage endolysin as antimicrobial agents to control SalmonellaTyphimurium.