Contemporary sugarcane cultivars originate from interspecific hybridization between Saccharum officinarum and Saccharum spontaneum, resulting in highly complex polyploid genomes. Dissecting chromosomal inheritance and structural variation is critical for accelerating molecular breeding in sugarcane. Here, we applied chromosome-specific painting using ten probes, together with S. spontaneum-specific probes, to resolve chromosomal composition and translocation patterns in the cultivar CT89-103 and its derived hybrids. High-resolution physical map was constructed in sugarcane cultivar CT89-103 that harbors 111 chromosomes, with 7-12 copies of each chromosome (1-10), and exhibits 13 distinct chromosomal translocation types. Its genome comprises 7.14% S. spontaneum-derived chromosomes and 24.11% interspecific recombinant chromosomes. The hybrids contain 108-116 chromosomes, with 5-15 copies of chromosomes 1-10 converging toward ~12 copies, and show similar genomic composition (8.20% S. spontaneum and 23.88% recombinant chromosomes). These findings demonstrate that S. spontaneum chromosomal segments and parental translocations are largely transmitted through n + n inheritance. Notably, extensive structural variation was observed, with 35 translocation types predominating in the hybrids. Altogether, this study provides a high-resolution view of chromosomal inheritance and structural variation in sugarcane, offering a cytogenetic framework for sugarcane hybridization.
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