Looking for the needle in a downsized haystack: Whole-exome sequencing unravels how selection and gene flow have shaped climatic adaptation in Douglas-fir (Pseudotsuga menziesii)

The widespread Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) occurs along a steep gradient of diverse climates throughout its natural range, which is expected to result in spatially varying selection to local climate conditions. However, phenotypic signals of climatic adaptation can often be confounded, because unraveled clines covary with signals caused by neutral evolutionary processes such as gene flow and genetic drift. Here, we present phenotypic and genotypic data from a common garden experiment showing a putative signal of adaptation to climate after trees have been growing for 40 years in a common environment. Sixteen Douglas-fir provenances originating from a North-to-South gradient of approx. 1,000 km were analyzed and genomic information was obtained from exome capture, which resulted in an initial genomic dataset of >90,000 single nucleotide polymorphisms. We used a restrictive and conservative filtering approach which permitted us to include only SNPs and individuals in environmental association analysis (EAA) that were free of potentially confounding effects (LD, relatedness among trees, heterozygosity deficiency and deviations from Hardy-Weinberg proportions). We used four conceptually different genome scan methods based on FST outlier detection and gene-environment association in order to disentangle truly adaptive SNPs from neutral SNPs and found that a relatively small proportion of the exome showed a truely adaptive signal (0.01-0.17%) when population substructuring and multiple testing was accounted for. Nevertheless, the unraveled SNP candidates showed significant relationship with climate at provenance origins which strongly suggests that they have most likely featured adaption in Douglas-fir across a steep climatic gradient. Two SNPs were independently found by three of the employed algorithms and one could be assigned with high probability to a Picea abies homolog gene involved in circadian clock control as was also found in Populus balsamifera.