The invention relates to a preparation and using method of a vibrio vulnificus rapid detection kit; wherein, the interior of the kit includes a loop-mediated isothermal amplification reaction liquid A, a Bst DNA polymerase B and a chromogenic agent C; the reaction liquid A contains a reaction buffer liquid, a dNTP, a magnesium sulfate, an upstream internal primer 5-ACCTGTGTTCCATGCTGCTTCTATTCGCAACTGACATGTCGG-3, a downstream internal primer 5-TAACAAAGGTCACCGTCCAGGCCAGAACGCAGGTCTTGTGAA-3, an upstream external primer 5-CCAGAGCCAGAGTTCTTCCT-3, a downstream external primer 5-GGCCCATCTCTTCCATCAC-3 and a betaine; the method for detecting the vibrio vulnificus includes the DNA extraction of the samples or the bacteria to be detected, the loop-mediated isothermal amplification reaction of the vibrio vulnificus and the chromogenic detection. The preparation and using method has the advantages of swiftness, strong specificity, high sensitivity and low cost.