Development of DNA markers that distinguish male and female haploid germlings of the brown alga, Cladosiphon okamuranus

Okinawa mozuku, Cladosiphon okamuranus, is one of many edible brown algae, yielding 22 Kton annually and contributing ¥5 billion to the Japanese economy. The life cycle of C. okamuranus is complicated, since the alga has self‐cloning life cycles in both haploid (N, male and female) and diploid (2N) conditions, but only diploid “seeds” (germlings) become edible sporophytes. Because haploid and diploid germlings are morphologically indistinguishable, haploid germlings are often mistakenly combined with diploid germlings for cultivation, which results in less efficient harvesting of mozuku. Sexual identification of haploid germlings is essential to develop better diploid strains by crossbreeding. With this aim, we performed RNA‐seq analysis of haploid germlings of C. okamuranus. Using its decoded diploid genome and transcriptomic information, we identified 269 genes that are expressed specifically in male or female haploids. BLAST analysis with Ectocarpus siliculosus gene models revealed that nine of 269 genes were putative sex determination‐related genes of C. okamuranus. A unique set of polymerase chain reaction primers for these nine genes was designed, and DNA amplification using primers enabled us to distinguish male and female haploid and diploid germlings. This tool will enable mozuku farmers to select diploid germlings free of haploid germlings. Using this DNA marker technique, the amount of mozuku cultivated in Okinawa is expected to increase.