Annotation and gene ontology analysis
To identify the potential role of significant SNPs in coding regions of
genes, genomic annotation was run using the blastn function
(Altschul et al., 1997) from BLAST (https://blast.ncbi.nlm.nih.gov/).
The 75 bp sequences associated with each SNP were annotated against theEucalyptus grandis genome (Myburg et al., 2014) and considered if
their significance valuesmet two related thresholds: an E ‐value
< 1 × 10−8 and a blast-score of at least
60.0. Chromosome number and location of significant SNPs were recorded,
as well as specific gene functions. The annotated SNPs were used to
predict broader biological functions using GO enrichment analysis
through the web interface PlantRegMap (Tian et al., 2020). GO terms are
organized within three categories: molecular function, cellular
component and biological process. We explored the biological process
aspect from the GO analysis, which refers to broad category of tasks
that the genes or gene products are programmed to achieve. Each of the
output GO terms delivered a set of genes that is associated with a
specific biological process. We use this output to explore how
functionally related SNPs, associated with the GO genes, might be
additively correlated with environmental factors (e.g., abiotic stress
response). For each climate variable, Fisher’s exact test was used to
test for significantly over-represented GO terms, with a threshold ofp-value <0.01. GO terms with the highest number of SNPs
and/or with pertinent biological
processes associated with environmental response (e.g., response to
heat, cold and drought) were recorded and considered for further
landscape genomic analysis.