Sequence Diversity of Co/Hd1 Homologs in Miscanthus sinensis

Miscanthus sinensis is a perennial C₄ grass native to eastern Asia, and a promising candidate bioenergy crop for cool temperate areas that are unsuitable for cultivation of its close relative, sugarcane. Flowering time is a crucial factor governing regional and seasonal adaptation of many plant species; additionally, it is also a key target trait for extending the vegetative phase of Miscanthus to improve biomass potential. To obtain insight into the control of flowering time for M. sinensis, we cloned homologs of Constans (Co)/Heading date 1(Hd1), which we call MsiHd1. From prior studies of other species such as Arabidopsis and rice, Co/Hd1 is known to be a key regulator of floral induction associated with photoperiod sensitivity. We compared the nucleotide sequence of MsiHd1 homologs among 18 wild-collected accessions of M. sinensis from Japan and 13 from China, by amplifying homolog-specific 2-2.5kb PCR products, followed by plasmid cloning then sequencing. Two to four MsiHd1 alleles in each accession were identified, suggesting that MsiHd1 consists of at least two loci in the Miscanthus genome. Verifying the open reading frame in MsiHd1, they were classified as putative functional alleles without mutations or non-functional alleles caused by insertions/deletions. Interestingly, only about half of all identified alleles in accessions from Japan were functional alleles (28 functional alleles/53 total alleles=52.8%), whereas accessions from China harbored mostly functional alleles (36 functional alleles/41 total alleles=87.8%). As ancestral populations of M. sinensis expanded their distribution from southern through northern Japan, selection for non-functional MsiHd1 alleles may have resulted in increased adaptation and fitness associated with earlier flowering, whereas in China a different adaptation strategy took place.