The invention relates to a positioning marking method of PGCs, and concretely relates to a positioning marking method for primordial germ cells (PGCs) of
scophthalmus maximus at embryonic development stage. The method comprises: acquiring
embryo samples of
scophthalmus maximus at all stages, fixing, using methanamide with the concentration of 50% to store the samples at -20 DEG C to 40 DEG C for usage; removing oolemma of all above
embryo samples of
scophthalmus maximus at all stages, using 1*PBST-
methanol eluants with different concentrations to perform gradient
dewatering processing, and storing at -20 DEG C to -40 DEG C; employing gradient
methanol o process the
embryo samples at all stages and subjected to above
dewatering processing for rewatering, then using a 1*PBST buffer to wash, and after eluting, pre-hybridizing at 60-65 DEG C for 1-2 h; after prehybridization, hybridizing in a hybridization liquid containing a scophthalmus maximus
RNA probe at 60-65 DEG C for a night; and after hybridization, washing, performing
antibody incubation and color development, and further performing positioning marking. The invention provides the relatively convenient and practical sample storage method, the problem that oolemma and
yolk are difficult to remove because
methanol dewatering and storage are performed during conventional embryo integral in-situ hybridization is solved, and a
protease K
digestion step is saved, and the operation steps are simplified.