The chromosome-level genome assembly of and comparative genomic analyses provide new resources and insights for understanding legume-rhizobial interactions.

The legume species (Chinese milk vetch [CMV]) has been widely cultivated for centuries in southern China as one of the most important green manures/cover crops for improving rice productivity and preventing soil degeneration. In this study, we generated the first chromosome-scale reference genome of CMV by combining PacBio and Illumina sequencing with high-throughput chromatin conformation capture (Hi-C) technology. The CMV genome was 595.52 Mb in length, with a contig N50 size of 1.50 Mb. Long terminal repeats (LTRs) had been amplified and contributed to genome size expansion in CMV. CMV has undergone two whole-genome duplication (WGD) events, and the genes retained after the WGD shared by Papilionoideae species shaped the rhizobial symbiosis and the hormonal regulation of nodulation. The chalcone synthase (CHS) gene family was expanded and was expressed primarily in the roots of CMV. Intriguingly, we found that resistance genes were more highly expressed in roots than in nodules of legume species, suggesting that their expression may be increased to bolster plant immunity in roots to cope with pathogen infection in legumes. Our work sheds light on the genetic basis of nodulation and symbiosis in CMV and provides a benchmark for accelerating genetic research and molecular breeding in the future.