Rapid sex identification method of spinach (Spinacia oleracea L.) in the vegetative stage using loop-mediated isothermal amplification

Main conclusion: A LAMP-mediated, simple and rapid method for sex identification in spinach was developed. Nutrient compositional analysis showed a higher iron content in male than female plants. Spinach (Spinacia oleracea L.) is a dioecious plant with its sex determined by the XY system. Male and female floral organs differ morphologically, but plants do not differ in the vegetative stage before flowering. PCR with Y chromosome markers has been used to determine the sex of dioecious plants before flowering. In this study, we developed a genotype-specific loop-mediated isothermal amplification (LAMP) for sex identification of individual vegetative-stage spinach plants, using primers designed for the genomic region flanked by male-specific markers. LAMP could specifically detect spinach males. The method was further modified to omit DNA purification and use just an aliquot of crude leaf extract homogenized in water. We compared the nutrient composition of males and females, finding higher amounts of iron in the males. Our method could therefore be used for rapidly discriminating male plants in the field, which is useful for efficient hybrid breeding.