Population Genetic Structure Analysis
Individuals named by species and location, we estimated the phylogenetic
relationships of
otherAquilegia species with the A. viridiflora complex to
determine whether the A. viridiflora complex in our study shared
an MRCA using IQ-TREE multicore version 1.6.12 (Nguyen, Schmidt, Von
Haeseler, & Minh, 2015) and MEGA X (Kumar, Stecher, Li, Knyaz, &
Tamura, 2018) with 1000 bootstrap replicates. Both the ML tree and NJ
tree indicated that 20 populations of the A. viridiflora complex
shared an MRCA. All trees were illustrated in iTOL (http://
itol.embl.de). Therefore, 672,439 high-quality SNPs in 20 populations of
the A. viridiflora complex were used for downstream analysis
after removing other Aquilegia species. To explore the patterns
of genetic structure of the A. viridiflora complex, we used a
phylogenetic network by the Neighbor-Net algorithm in the software
Splits Tree (Huson & Bryant, 2006) with 1000 bootstrap replicates.
ADMIXTURE v.1.3.0 (Alexander, Novembre, & Lange, 2009) was applied to
investigate the maximum likelihood of the ancestry of all individuals
with K values ranging from 2 to 10 with 10 replicates for each K value
and examined the optimum K value according to the lowest value of the
error rate. Principal component analysis (PCA) was performed using
EIGENSOFT v.6.1.4 (Price et al., 2006) to infer population genetic
structure. By combining the results of the phylogenetic relationship and
genetic structure analysis to establish lineages for downstream
analysis, we divided 20 populations into four lineages. Since the
results of the population genetic structure showed a mixed genetic
background in some individuals, the Python package HyDe was used to
identify hybridization events at the individual level (Blischak,
Chifman, Wolfe, & Kubatko, 2018). Among them, P2 was an individual who
did not show a mixed genetic background, and P1 and P3 were individuals
of other populations, respectively.