Alfalfa (Medicago sativa) is one of the most important forage crops in the world; however, its molecular genetics and breeding research are hindered due to the lack of a high-quality reference genome. Here, we report a de novo assembled 816-Mb high-quality, chromosome-level haploid genome sequence for ‘Zhongmu No.1’ alfalfa, a heterozygous autotetraploid. The contig N50 is 3.92 Mb, and 49 165 genes are annotated in the genome. The alfalfa genome is estimated to have diverged from M. truncatula approximately 8 million years ago. Genomic population analysis of 162 alfalfa accessions revealed high genetic diversity, weak population structure, and extensive gene flow from wild to cultivated alfalfa. Genome-wide association studies identified many candidate genes associated with important agronomic traits. Furthermore, we showed that MsFTa2, a Flowering Locus T homolog, whose expression is upregulated in salt-resistant germplasms, may be associated with fall dormancy and salt resistance. Taken together, these genomic resources will facilitate alfalfa genetic research and agronomic improvement.

苜蓿（苜蓿）是世界上最重要的牧草作物之一。然而，由于缺乏高质量的参考基因组，其分子遗传学和育种研究受到阻碍。在这里，我们报道了杂种的自体四倍体“中母一号”苜蓿的从头组装816-Mb高质量，染色体水平的单倍体基因组序列。重叠群N50为3.92 Mb，基因组中标注了49 165个基因。苜蓿基因组被估计从发散蒺藜苜蓿大约八百万年前。对162种苜蓿种质进行的基因组种群分析显示，遗传多样性高，种群结构弱，并且从野生到栽培苜蓿的基因流广泛。全基因组关联研究确定了许多与重要农艺性状有关的候选基因。此外，我们显示MsFTa2，一个开花的基因座T同源物，其表达在耐盐种质中上调，可能与秋季休眠和耐盐性有关。这些基因组资源合在一起将促进苜蓿的遗传研究和农艺学的改进。