Bacteriophages have been intensively-studied and applied in a variety of practical applications such as biocontrol of food-borne pathogens. Especially, endolysin of bacteriophage is highly specific to host strains that can hydrolyze cell wall. To get the control agent for foodpoisoning Bacillus cereus, cloning and nucleotide sequence of putative endolysin from temperate phage 250 (Tp250) of emetic B. cereus 250 were investigated. Tp250 was induced by mitomycin C and belonged to Siphoviridae family. Tp250, a circular dsDNA genome of 56,505 bp, consisted of various structural and functional genes for survival or propagation. ORFs from Tp250 were involved in replication, head morphogenesis, head-to-tail joining, tail morphogenesis, lysogenic module, host lysis, and other functional genes such as related metabolisms. For cloning of putative endolysin, Tp 250 DNA was extracted and pLYS250 was constructed. The pLYS250 had 752 ORF encoding a putative endolysin with 250 amino acids, which was expressed by around 28 kDa in Escherichia coli. Therefore, Tp250 and the endolysin might be applied as one of agents to reduce B.cereus pathogen in foods directly or through over-expression and purification.