Transcriptome analysis of alfalfa (Medicago sativa L.) roots reveals overwintering changes in different varieties

Low temperatures are one of the major abiotic stresses that affect alfalfa's development and yield. Enhancing frost resistance through resistance-related genes is one of the most effective ways to address this issue in alfalfa. Therefore, exploring cold-resistant gene resources and the cultivation of cold-resistant alfalfa cultivars is inevitable in order to achieve high yield and quality. In this study, we conducted transcriptome profiling of roots obtained from two alfalfa genotypes, i.e., Qingda No.1 for freeze tolerance and Gannong No.9 for freeze sensitivity. We observed that Qingda No.1 had more lateral roots and a more developed root system after overwintering, while Gannong No.9 had fewer lateral roots and an underdeveloped root system. After overwintering, Qingda No.1 exhibited higher superoxide dismutase (SOD) activity compared to Gannong No.9, while Gannong No.9 showed higher perosuperoxide dismutasexidase (POD) activity than Qingda No.1. We identified 25,935 differentially expressed genes, with 12 979 and 12 956 differential genes found in the freeze-tolerant variety Qingda No.1 group and the freeze-sensitive Gannong No.9 group, respectively. The enrichment of the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways also differed between the two groups. We also discovered several gene family members, and the most frequent transcription factors were bHLH, B3, NAC, WRKY, and MYB_related. These findings provide comprehensive information to further understand the molecular mechanisms of adaptation to freezing stress in alfalfa and offer potential functional candidate genes for adaptation to abiotic stress.