Abstract
While water content is a critical factor affecting the outcome of cryopreservation, the mechanism by which it influences seed viability after cryopreservation remains unclear. In this study, Paeonia lactiflora seeds with different water content as experimental materials, analyzed the seed viability, oxidative stress indicators, and transcriptomics before and after cryopreservation, to explore the mechanism of the seed viability changes. The results demonstrated that after cryopreservation, seeds with high water content displayed reduced viability, along with abnormal accumulation of reactive oxygen species (ROS) content, which subsequently triggered ROS-mediated oxidative damage. In contrast, seeds with low water exhibited enhanced following cryopreservation, their ROS levels remained relatively stable, while notable alterations were detected in multiple antioxidant system parameters. At the transcriptional level, 1224 differentially expressed genes (DEGs) up-regulated and 1839 DEGs were down-regulated in seeds with high water content after cryopreservation, but 2090 DEGs up-regulated and 1783 DEGs down-regulated in the seeds with low water content. Among them, 687 DEGs were common to both the high- and low-water content seed groups. Gene Ontology (GO) functional analysis indicated that these DEGs are primarily involved physiological metabolic processes including osmotic response, glucosidase activity, protein kinase binding, and response to hydrogen peroxide. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis confirmed that the Mitogen-Activated Protein Kinase (MAKP) plant signaling pathway and the starch and sucrose metabolism pathway are the key pathways governing the response of seeds with varying water contents to cryopreservation. Finally, through weighted gene co-expression network pinpointed DHN1 and LEA34 as the core genes regulating changes in seed viability after cryopreservation. These findings offer a broader perspective for in-depth exploration of the molecular regulatory mechanisms underlying the differences in seed viability changes after cryopreservation and are crucial for comprehensively clarifying the precise pathways via which these key genes regulate seed viability changes after cryopreservation.