Genome-Wide Association Study (GWAS) for Freezing and De-Acclimation Tolerance in Polish Winter Barley

Winter survival in barley depends on freezing tolerance and de-acclimation tolerance, yet their genetic determinants under increasingly unstable winters remain poorly understood. Here, 188 Polish barley accessions were evaluated over two consecutive growing seasons (2021–2022) using genome-wide association studies (GWAS) with a mixed-linear model (MLM) and high-density single nucleotide polymorphism (SNP) and diversity arrays technology sequencing (DArTseq) markers. Freezing and de-acclimation tolerance were quantified by 16 chlorophyll fluorescence parameters and post-freezing survival rates in plants subjected to 21 days of cold acclimation (4 °C/2 °C, day/night) and 7 days of de-acclimation (12 °C/5 °C, day/night). The results showed that freezing and de-acclimation tolerance are related but genetically distinct. The cold-acclimated (CA) state exhibited significant marker–trait associations on chromosomes 2H and 6H, whereas the de-acclimated (DA) state displayed a broader, more complex genetic architecture, particularly on chromosomes 2H and 7H. Fv/Fm showed the strongest associations for both SNP and DArTseq markers in both states. PI(csm), followed by PI(cs0) and PI(total), showed high SNP associations in the DA state, indicating a strong relationship between photosynthetic performance and freezing tolerance after de-acclimation. Notably, the DArTseq marker 11400277 on chromosome 7H showed multiple marker–trait associations across both states. These findings provide a genomic basis for marker-assisted selection of climate-resilient winter barley cultivars.