The invention relates to the field of molecular genetics and in particular relates to a molecular marking method of a pig USP (Universal Stress Protein) 18 gene 5' regulation region mutation site and application of the molecular marking method in pig blue-ear disease resistant breeding. The inventor finds that USP185' regulation regions of dapulian pigs and dudachang hybrid pigs have a plurality of differences, wherein the -1533 site has A to G mutations; according to a luciferase reporter gene system, after a fixed point A of the -1533 site is mutated into G, the promoter activity of the -1533 site is remarkably reduced; the A allelomorphic gene frequency of the -1533 site and the blue-ear disease resistant duplian pigs is high; and the mRNA expression level of the USP18 gene is high and consistent. The genetype of the -1533 site of the USP18 gene regulation region in the pig genome is detected as the molecule marker associated with the pig blue-ear disease resistant characteristics, so that not only can the method be simple and quick, but also the method is not affected by the environment, and therefore, early breeding selection can be realized.