Development and interlaboratory validation of a cultivar-specific identification method for the table grape ‘Shine Muscat’ using loop-mediated isothermal amplification (LAMP)

Abstract

‘Shine Muscat’ is an elite table grape cultivar developed by the National Agriculture and Food Research Organization in Japan. Recently, the infringement of breeders’ rights in various fruits has become a serious problem in Japan. In this study, a loop-mediated isothermal amplification (LAMP)-mediated cultivar identification method for ‘Shine Muscat’ was developed. We comprehensively analyzed retrotransposon insertion sites using 24 major grape cultivars and identified two insertions, VINE1-Cl160 and VINE1-Cl155, which are unique to ‘Shine Muscat’. LAMP primers targeting VINE1-Cl160 and VINE1-Cl155 were designed, and specific amplifications were confirmed. We also designed a primer set to detect the grape endogenous reference sequence, UDP-glucose:flavonoid 3-O-glucosyltransferase. To improve rapidness and cost-effectiveness, we applied single-stranded tag hybridization on a chromatography printed-array strip system, a lateral flow DNA chromatography technology. The developed method was validated with an interlaboratory study. This novel identification method would be particularly useful for border inspections.