Sammendrag
Scots pine (Pinus sylvestris L.) is the most widely distributed pine species across Eurasia and is a vital genetic resource in Norway with a widespread geographic range. It is one of the most suitable and best pioneer species for the afforestation because of its undemanding nature and higher adaptability potential in poor soil conditions. This study investigated population structure, GWAS (Genome wide association studies), and Gene ontology using 50K SNP genotyping data as well as DNA methylation analyses from reduced representation bisulfite sequencing (RRBS) data. Population structure was estimated using fastSTRUCTURE, STRUCTURE 2.3.4 and STRUCTURE HARVESTER, GWAS was performed using Plink v2.0, gene ontology through R-stats and DNA methylation through RefFreeDMA (Reference free DNA methylation analysis) software, further differentially methylated regions functionally characterized via BLAST algorithms using QIAGEN CLC Genomic Workbench 22.0.2. Population structure analysis of 310 individual samples identified the most likely K-value of 8, and genetic diversity within all the cluster showed significant variability; mean FST values indicated genetic differentiation, particularly for clusters 5 and 8. Cluster 4 was the largest with 240 of 310 trees and was therefore presumed to represent trees originating from the sowed Swedish seed. GWAS identified distinct SNP markers associated with plant height (PH) based on different allele and genotype models, with markers in the dominant allele model predominating. Interaction analysis further explained allele-specific associations particularly observed in Swedish trees across the forest plots. Analyses were expanded to investigate influence of seasonal variation in annual growth of plants by correlating SNP markers, growth associated SNPs varied significantly across seasons; dominant allele models showed highest associations during periods of High SPEI. Gene ontology (GO) analysis provided insights into biological importance of identified genes, identifying different molecular functions, biological processes, and cellular components associated with PH. Particularly, a subset of genes showed shared and unique functions across different samples and allele models, emphasizing detailed genetic mechanisms underlying PH and plant growth. RRBS analyses of 29 sampled trees, roughly 50:50 Swedish:Norwegian and 50:50 tall:short, identified 394 differentially methylated fragments between Norwegian and Swedish seed origin, 129 in CpG, 128 in CHG, and 80 in CHH. Norwegian trees showed higher prevalence of methylation in CHH site, while Swedish trees showed higher methylation in CpG. In the tree height comparison, 593 DMRs were identified between tall and short trees: 293 in CpG, 188 in CHG, and 112 in CHH. Individuals in both tree growth categories (tall and short) exhibited high methylated sites in CHH, while fewer methylated sites were observed in CHG and CpG for tall trees and short trees, respectively. Therefore, our detailed analysis enhances the understanding of underlying genetic and epigenetic mechanisms regulating Scots pine growth traits. And the findings of this study provide valuable insights for conservation practices and sustainable forest management as well as breeding approaches directed towards enhancing resilience and productivity in Pinus sylvestris.
Keywords: Scots Pine; Pinus sylvestris; SNP; population structure; GWAS; gene ontology; DNA methylation