FIGURE 7 Phylogenetic tree constructed with the whole sequences of the
chloroplast genomes of the above 59 species.
3.6 | Selective pressure
analysis
We calculated the Ka/Ks values of Polygonatum protein-coding
genes (PCGs) based on KaKs_Calculator2. The genes subject to positive
selection are matK , ndhA , petB , and ycf2 ,
with a maximum Ka/Ks value of 1.41. The genes that were positively
selected can be categorized into five groups according to their
functions: (1) maturation enzyme gene matK ; (2) photosynthetic
system genes ndhA and petB ; and (4) the ycf2 gene
with unknown function. The remaining genes, all with Ka/Ks values less
than 1, were subjected to purifying selection, with atpI being
the lowest at 0.048 (Figure S3).
3.7 | Phylogenetic
analysis
Phylogenetic trees were constructed using the maximum likelihood (ML)
approach based on 59 complete chloroplast genome sequences ofPolygonatum medicinal plants published in the NCBI database
(Dioscorea aspersa and Dioscorea alata ) to illustrate the
genetic relationships of these species. The results showed that the two
outgroups were distinct from Polygonatum . The nodes of the
phylogenetic tree in this study had a high support rate and strong
reliability for the phylogenetic analysis (Figure 7, Table S15). In the
phylogenetic trees, all alternate leaf types of Polygonatum species were clustered on one large clade and species with verticillate
leaves were divided into sect. Sibirica and sect. Verticillata group, confirming that the leaf type affects the
classification of the genus. Of these, sect. Polygonatum and sect. Sibirica clustered together and exhibited a sister
relationship with sect. Verticillata. Among the sixPolygonatum species, P. sibiricum formed a monophyletic
group (sect. Sibirica ) with 100 % support. P.
cyrtonema was the earliest clade to differentiate, followed P.
odoratum , P. filipes and other related species clustered into a
sister clade in sect. Polygonatum . P. kingianum was the earliest species differentiated from sect. Verticillata. P. zanlanscianense and other verticillate
leaf type species of Polygonatum clustered into sister clades
with P. kingianum. These results are consistent with those of the
previous studies.
In addition, we downloaded the chloroplast genomes of six otherPolygonatum species from the database of the National Center for
Biotechnology Information (NCBI) to explore the chloroplast genome
affinities of Polygonatum in different regions. The phylogenetic
results showed that P. cyrtonema was collected in Anhui Province
and had the closest relationship with P. cyrtonema (MZ579646,
MZ150839) stemming from Zhejiang Province and a relatively distant
relationship with Hunan (OL436258), Beijing (MW248135), and Jiangxi
(MZ029094). P. zanlanscianense (from Yunnan Province) was closely
related to Guizhou Province (MW373522), followed by Hubei Province
(OL405020) and Hunan Province (ON534059).