Sugarcane is among the most efficient crops in converting solar energy into chemical energy. However, due to its complex genome structure and inheritance, the genetic and molecular basis of biomass yield in sugarcane is still largely unknown. We created an F2 segregating population by crossing S. officinarum and S. spontaneum and evaluated the biomass yield of the F2 individuals. The F2 individuals exhibited clear transgressive segregation in biomass yield. We sequenced transcriptomes of source and sink tissues from 12 selected extreme segregants to explore the molecular basis of high biomass yield for future breeding of high-yielding energy canes. Among the 103,664 assembled unigenes, 10,115 and 728 showed significant differential expression patterns between the two extreme segregating groups in the top visible dewlap leaf and the 9 culm internode, respectively. The most enriched functional categories were photosynthesis and fermentation in the high-biomass and the low-biomass groups, respectively. Our results revealed that high-biomass yield was mainly determined by assimilation of carbon in source tissues. The high-level expression of fermentative genes in the low-biomass group was likely induced by their low-energy status. Group-specific expression alleles which can be applied in the development of new high-yielding energy cane varieties via molecular breeding were identified.