Salmonella is one of the leading foodborne pathogens responsible for illnesses and hospitalizations, and responsible for 38% of the 3000 deaths reported annually (US Center for Disease and Control). Salmonella, present in the gastrointestinal tract of poultry, is passed from bird to bird for instance through contaminated feed and water, environmental sources/vectors, etc. Bacteriophages are bacterial viruses and therefore natural predators of bacteria. Bacteriophages (phages) are harmless to humans and animals, and have been extensively used for prevention and control of bacterial pathogens. In vitro experiments show that phages, alone and in cocktail, can decrease significantly Salmonella levels in contaminated foods; however, the future of phages in food safety is further dependent on the regulatory agencies that still display uneasiness when it comes to the use of phages. This is mostly due to a scarcity of strong scientific evidence, emergence of phage resistant bacterial phenotypes, and the high percentage of genes in phages with unknown function, which can be virulence factors or toxin genes. The main objective of this work was to address the problem of genes of unknown function in a phage genome. To achieve this, the genome sequence of a phage was analysed using bioinformatics tools, all unknown genes were identified and later the whole genome of the phage will be introduced in a bacterial artificial chromosome so that it could be genetically engineered. Phage vB_SenS_Φ38, targeting Salmonella enterica serovar Enteritidis, was genetically characterized and was shown to be 42439 bp in size, dsDNA and to contain 60 open reading frames (ORFs). The study further focused in the 28 hypothetical proteins that exist in the phage genome which were effectively cloned and their antimicrobial activity evaluated. These 28 hypothetical proteins have not shown any antibacterial function, either when expressed in liquid or in solid medium.