Abstract:The genome DNA of Staphylococcus aureus was extracted by four different method and finally chose a method combined steel ball destruction and acetone，protease K removing impurities to get high purity and output DNA.Then the protein gene（egc）of Staphylococcus aureus，a NA gene of Salmonella，and ipaH gene （iapH）of Shigella were used as the gene targets.And the samples were determined by real-time PCR technique.The results showed that the primers were highly specific.No fluorescence signal was determined in other 3 control pathogens（Listeria monocytogenes，Vibrio parahaemolyticus and Bacillus cereus）but target bacteria. The RT-PCR assay could be specific and rapid，and the detection limits were 10-6，10-6，10-7ng/μL.