URANO, Akihisa*; MORIYA, Shogo; ABE, Shuiti; Hokkaido Univ, Sapporo; Res Dev Ctr, Nisshinbo, Chiba; Hokkaido Univ, Hakodate: Oligonuclotide DNA Microarray for Haplotype Analysis in Field Endocrinology

Wild animals of the same species often form a mixed population of individuals from different stocks. Analyses of endocrine parameters, such as the contents of hormones and their plasma levels, in individuals require identification of their genetic stocks, otherwise mislead incorrect conclusions. Oceanic salmon in the Bering Sea contain large mixed populations, in which origins of fish varied from Asia to North America. Identification of their natal country is thus crucial for understanding of endocrine mechanisms of their migratory behavior. We therefore developed a method in which oligonucleotide DNA microarray is used for detection of nucleotide sequence variations in the 5� half of the mitochondrial DNA control region of chum salmon. The method includes 1) immobilization of 17 to 20 mer synthetic oligonucleotides, which contain the previously identified polymorphic site sequence, on a slide glass pre-coated with poly-carbodiimide resin, 2) hybridization of biotinylated PCR products spanning the 5� variable portion with a DNA microarray, and 3) coloring of hybridization signals by a conventional ABC method, and comparison of scanned signal images on a computer. All processes of hybridization and detection could be completed within eight hours, so that we obtained the result even during a few week research cruise. Since the present oligonucleotide microarray method can be carried out without any specialized laboratory equipment, it will be a rapid and accurate means for genetic stock identification of animals in the field.