Abstract: To establish a new rapid and accurate method to detect Vibrio parahaemolyticus,principle of rolling circle amplification (RCA) technique was used to design primers,padlock probe and reaction system.The padlock probe consists of two target complementary regions at 5'and 3'end,it will form a ring under the function of DNA ligase,then use probe rolling circle to amplify nucleotide and verify products with gel electrophoresis.The result showed that the method can detect Vibrio parahaemolyticus specifically,and the detection sensitivity of RCA was higher than that of conventional PCR.It proved that RCA is a capable approach to detect Vibrio parahaemolyticus.The method is specific,stable and reliable.