Woody bamboos (Bambusoideae) are renowned for its polyploidy and rare flowering. Bambusa odashimae is one of the bamboo species with the highest chromosome count (104) in the subfamily and has the highest heterozygosity of all sequenced bamboo genomes so far. Compared with other bamboo species, it can efficiently utilize exogenous hormones to regulate in vitro flowering, providing valuable insights into the hormonal regulation of bamboo flowering. Here, we generated the haplotype-resolved genome assembly of B. odashimae, despite the complexity and high chromosome number, supplemented by thirty-three transcriptomes from eleven developmental periods using a tissue culture system. The assembled genome can be divided into Haplotype I, Haplotype II, and Haplotype III, each containing A, B, and C subgenomes. Haplotype I may be derived from Dendrocalamus whereas Haplotypes II and III are closely related to Bambusa, indicating that B. odashimae has an origin involving both intergeneric and interspecific hybridizations. The high heterozygosity renders the possibility to detect abundant allele-specific expression (ASE), with ASE genes enriched in cytokinin-related pathways, likely associated with efficient cytokinin-promoted flowering. Notably, we found that the CONSTANS (CO) genes were potentially key regulators of in vitro flowering in B. odashimae. Overall, based on the in vitro system combined with a high-quality reference genome, our study provides critical insights into the origin of this nonaploid bamboo and links hybridization and in vitro flowering in bamboos.

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