Sample collection of Allium macrostemon
We collected bulbs from 47 A. macrostemon plants from different locations in Japan, including fields and riverbanks. Figure 1 and Table 1 show details regarding the plants, their collection dates, and places of origin. We further created three technical replicate samples, resulting in a total of 50 samples subjected to ddRAD-Seq.
Table 1
Information on Allium macrostemon samples used in the study.
No | Isolate | Location (City, district, prefecture) | Collection date | Group |
5 | WOA360 | Inakadate-mura, Minamitsugaru-gun, Aomori | 8-Aug-14 | Group C1 |
33 | WOAK354 | Shincho, Yonezawa, Akita | 15-Jun-14 | Group C1 |
22 | WOYA331 | Shiromori, Yamagata, Yamagata | 12-Jun-14 | Group C1 |
10 | WOYA335 | Kawatoi, Nanyo, Yamagata | 12-Jun-14 | Group C1 |
24 | WOFK488 | Akogashima, Atami-machi, Koriyama, Fukushima | 23-Apr-15 | Group C2 |
21 | WOMY326 | Nishiutari, Hamaichi, Higashimatsushima, Miyagi | 13-Jun-14 | Group B |
2 | WOGU466 | Shimotoyooka-machi, Takasaki, Gunma | 16-Mar-15 | Group B |
12 | WOIB449 | Senba-cho, Mito, Ibaraki | 15-Mar-15 | Group B |
30 | WOTK389 | Utsuki-machi, Hachioji, Tokyo | 7-Dec-14 | Group B |
31 | WOTK389_1 | Utsuki-machi, Hachioji, Tokyo | 7-Dec-14 | Group B |
48 | WOC441 | Kamezawa, Futtsu, Chiba | 14-Mar-15 | Group C1 |
29 | WOKN373 | Terasaka, Oiso-machi, Naka-gun, Kanagawa | 5-Dec-14 | Group C4 |
25 | WOST86 | Kawaguchi, Saitama | 8-Jul-13 | Group B |
11 | WONI551 | Nishinaka, Itoigawa, Niigata | 18-Apr-15 | Group C2 |
7 | WOTY540 | Shiragawa, Himi, Toyama | 18-Apr-15 | Group C2 |
23 | WOIS532 | Shima-machi, Komatsu, Ishikawa | 17-Apr-15 | Group C2 |
18 | WOFI554 | Kumasaka, Awara, Fukui | 17-Apr-15 | Group C3 |
49 | WOAI220 | Gonjochi62, Tahara, Aichi | 18-Apr-14 | Group C1 |
8 | WOSH405 | Echigoshima, Yaizu, Shizuoka | 24-Jan-15 | Group B |
6 | WOSG538 | Yasu, Shiga | 16-Apr-15 | Group C4 |
1 | WOKY536 | Shimogyo-ku, Kyoto, Kyoto | 17-Apr-15 | Group C3 |
47 | WOHG425 | Shimizuueno, Uozumi-machi, Akasi, Hyogo | 7-Mar-15 | Group C4 |
9 | WONR546 | Denen, Gozyo, Nara | 20-Apr-15 | Group C1 |
27 | WOME413 | Koazakacho, Matsusaka, Mie | 25-Jan-15 | Group B |
13 | WOW560 | Wakayama, Wakayama | 20-Apr-15 | Group C4 |
19 | WOT105 | Hoki-cho, Saihaku-gun, Tottori | 14-Nov-13 | Group C4 |
20 | WOOY574 | Tamashimakurosaki, Kurashiki, Okayama | 9-Mar-15 | Group C3 |
42 | WOOY575 | Okutyofukutani, Setouchi, Okayama | 8-Mar-15 | Group A |
35 | WOHR72 | Yahatahigashi, Nakaji, Itsukaichi-cho, Saeki-ku, Hiroshima, Hiroshima | 10-Mar-13 | Group A |
26 | WOKW581 | Konanchooka, Takamatsu, Kagawa | 10-Mar-15 | Group A |
34 | WOKW582 | Ayautachookadakami, Marugame, Kagawa | 10-Mar-15 | Group A |
3 | WOTS607 | Kamo, Higashimiyoshi-cho, Miyoshi-gun, Tokushima | 10-Mar-15 | Group C3 |
4 | WOEH562 | Niya, Imabari, Ehime | 13-Mar-15 | Group C1 |
36 | WOEH566 | Takata, Tsushima-cho, Uwajima, Ehime | 13-Mar-15 | Group A |
14 | WOKO594 | Sakihama-cho, Muroto, Kochi | 12-Mar-15 | Group B |
15 | WOKO594_1 | Sakihama-cho, Muroto, Kochi | 12-Mar-15 | Group B |
39 | WOF366 | Kitano-machi, Kurume, Fukuoka | 14-Nov-14 | Group A |
40 | WOF395 | Higashi-ku, Fukuoka, Fukuoka | 16-Dec-14 | Group C4 |
43 | WOS317 | Saga, Saga | 13-Feb-15 | Group C1 |
28 | WOO199 | Akimachishimobaru, Kunisaki, Oita | 12-Feb-15 | Group C4 |
45 | WOO200 | Ishigakihigashi, Beppu, Oita | 12-Feb-15 | Group C3 |
50 | WON227 | Shirakami, Unzen, Nagasaki | 9-Feb-15 | Group C4 |
41 | WOK289 | Shimoshima, Amakusa, Kumamoto, | 9-Feb-15 | Group C3 |
38 | WOM366 | Komono, Hanagashima-cho, Miyazaki, Miyazaki | 11-Feb-15 | Group C4 |
46 | WOM367 | Noziri-cho, Kobayashi, Miyazaki | 11-Feb-15 | Group C3 |
44 | WOM368 | Sakataniko, Nichinan, Miyazaki | 12-Feb-15 | Group C1 |
37 | WOKG194 | Hamabira, Tarumizu, Kagoshima | 9-Feb-15 | Group C3 |
16 | WOKG195 | Myoken-machi, Makurazaki, Kagoshima | 10-Feb-15 | Group C3 |
17 | WOOK122 | Chunjun, Kitanakagusukuson, Nakagami-gun, Okinawa | 10-Dec-13 | Group C4 |
32 | WOOK122_1 | Chunjun, Kitanakagusukuson, Nakagami-gun, Okinawa | 10-Dec-13 | Group C4 |
Variants detection using RAD-Seq data
The ddRAD-Seq of the 50 samples (Table 1) generated more than 10 Gbp of data, comprising a total of 196.2 million raw, single-end 51-bp reads. Quality-based filtering yielded an average of 0.9 million reads (ranging from a minimum of 0.2 million to a maximum of 3 million) across the 50 samples (Supplementary Table S1). The Stacks program-built loci de novo with an average coverage depth of 16.78-fold (Supplementary Table S2). We identified a total of 5,848 variant sites for subsequent analysis.
The plant consists of three groups
Principal component analysis (PCA) of these variant sites effectively partitioned the 50 samples into three groups, labeled A, B, and C (Fig. 2). The division into these three groups was primarily driven by the principal components 1 and 2, which contribute 16.5% and 9.86% respectively. Analysis of principal components 3, 4, and 5 did not reveal additional meaningful features (Supplementary Fig.S1). Multidimensional scaling (MDS) analysis classified the samples into three groups, with the same members as identified by the PCA (Fig. 3).
Members of Group A originated from three districts in Japan: the plants in Chugoku (35 and 42), Shikoku (26, 34, and 36), and Kyushu (39). Thus, they were found throughout Western Japan, excluding the Okinawa district. Members of Group B, on the other hand, came from five districts in Japan: plants from Tohoku (21), Kanto (2, 12, 25, and 30, with 30's technical replicate 31), Chubu (8), Kinki (27), and Shikoku (14, and its technical replicate 15). These were not present in the Chugoku, Kyushu, and Okinawa districts, indicating that they were not found in Western Japan without a presence in the Shikoku district. Members of Group C were from all districts in Japan: plants in Tohoku (5, 10, 22, 24, and 33), Kanto (29 and 48), Chubu (7, 11, 18, 23, and 49), Kinki (1, 6, 9, 13, and 47), Chugoku (19 and 20), Shikoku (3 and 36), Kyushu (16, 28, 37, 38, 40, 41, 43, 44, 45, 46, and 50), and Okinawa (17 and its technical replicate 32).
Cluster analysis was performed using a pairwise distance matrix (Fig. 4), which identified the same three groups as those identified in the PCA and MDS analyses: groups A and B each formed a tight cluster, while group C was an outgroup of groups A and B. The cluster analysis detected several subclusters, but, with two exceptions, it did not detect any region-specific subclusters. The two exceptions are the subclusters to which plants 5, 10, 22, and 33 from the Tohoku district and plants 16 and 46 from the Kyushu district belong.
Gene flow from group A and/or group B to group C was detected, but not from group C to group A or group B
We performed an admixture analysis to estimate the degree of genetic mixing. We used values from 1 to 10 as possible K values (symbolizing the number of ancestral populations), computed the cross-validation (CV) errors, and estimated the most likely K value, i.e., the K value with the lowest CV (Supplementary Fig. S2). The CV error was minimized at K = 3, indicating that the most likely number of ancestral populations is three, aligning with the number of groups identified above. The admixture analysis results are displayed (Fig. 5, Supplementary Fig.S3). The admixture analysis detected no gene flow into groups A and B from other groups. Conversely, this analysis identified gene flow into group C from either group A, group B, or both.
Group C consists of four subgroups
Analysis of the results at K = 3, the highest possible value, divided the individuals into four subgroups: subgroup C1, with no detected gene flow from either group A or group B (plants 4, 5, 9, 10, 22, 33, 43, 44, 48, and 49); subgroup C2, with detected gene flow only from group A (plants 7, 11, 23, and 24); subgroup C3, with detected gene flow only from group B (plants 1, 3, 16, 18, 20, 37, 41, 45, and 46); and subgroup C4, with detected gene flow from both groups A and B (plants 6, 13, 17 (and its technical replicate 32), 19, 28, 29, 38, 40, 47, and 50). Subgroups C1, C2, and C3 were also found in the PCA and MDS analyses. Cluster analysis demonstrated that subgroups C1, C2, and C3 formed one or two clusters. Additionally, subgroup C1 was the most distant from groups A and B in the PCA and MDS analyses. We found no specific characteristics tied to regions within each subgroup. The admixture analysis also displayed more intense gene flow into group C from group B than from group A.
Statistical interpretations of the classifications
We performed a statistical analysis among the six populations. The Fst value analysis (Table 2) exposed a high level of genetic differentiation between groups A and B, yet a comparatively low average degree of genetic differentiation between group A and subgroups C1, C2, C3, or C4, as well as between group B and any of these subgroups. Notably, the genetic differentiation between group B and either C1 or C2 was higher than that between group A and either C1 or C2. Conversely, the Fst values between group B and either C3 or C4 were lower compared to those between group A and either C3 or C4.
Table 2
Fst values among the six groups.
| Group B | Group C1 | Group C2 | Group C3 | Group C4 |
Group A | 0.300503 | 0.159082 | 0.176953 | 0.163133 | 0.124971 |
Group B | | 0.170023 | 0.211171 | 0.070683 | 0.076914 |
Subgroup C1 | | | 0.059202 | 0.062329 | 0.053722 |
Subgroup C2 | | | | 0.089261 | 0.053697 |
Subgroup C3 | | | | | 0.021505 |
For each population, we conducted a statistical analysis. The average values of nucleotide diversity (π) and mean expected heterozygosity (He) were higher in the subgroups of group C; these values in groups A and B were similar and lower than in group C (Table 3, Supplementary Table S3). A negative Fis value indicates avoidance of inbreeding in groups A, B, and C2, while the relatively high Fis values in groups C1, C3, and C4 imply a moderate degree of inbreeding (Table 4).
Table 3
Population genetic statistics of each group.
| Mean expected heterozygosity (He) | Mean value of π | Mean measure of Fis |
Group A | 0.14653 | 0.16737 | -0.10965 |
Group B | 0.15041 | 0.16267 | -0.14291 |
Group C1 | 0.20474 | 0.22111 | 0.04932 |
Group C2 | 0.15921 | 0.19686 | -0.04108 |
Group C3 | 0.21477 | 0.23195 | 0.0379 |
Group C4 | 0.22037 | 0.23575 | 0.12327 |
Table 4
Conservation of heterozygous loci positions across pairs of samples in each group. Some pairs were selected and shown. Full data are presented in Supplementary Table S5.
Group | Plant | Plant | Number of variable sites | Number of conserved heterozygous sites | % |
A | No.26 (Shikoku) | No.39 (Kyushu) | 12344 | 5574 | 45.1 |
No.34 (Shikoku) | No.36 (Shikoku) | 15116 | 6166 | 40.8 |
No.35 (Chugoku) | No.39 (Kyushu) | 11135 | 4961 | 44.5 |
No.35 (Chugoku) | No.42 (Chugoku) | 17896 | 6545 | 36.6 |
No.39 (Kyushu) | No.34 (Shikoku) | 9726 | 5065 | 52.1 |
No.39 (Kyushu) | No.35 (Chugoku) | 11015 | 5078 | 46.1 |
No.42 (Chugoku) | No.34 (Shikoku) | 14962 | 5987 | 40.0 |
B | No.2 (Kanto) | No.8 (Chubu) | 6160 | 2810 | 45.6 |
No.2 (Kanto) | No.21 (Tohoku) | 9316 | 3695 | 39.6 |
No.8 (Chubu) | No.12 (Kanto) | 10640 | 4336 | 40.7 |
No.8 (Chubu) | No.27 (Kinki) | 7444 | 3702 | 49.7 |
No.12 (Kanto) | No.14 (Shikoku) | 22672 | 10740 | 47.4 |
No.14 (Shikoku) | No.15 (Shikoku) | 16321 | 7997 | 49.0 |
No.30 (Kanto) | No.31 (Kanto) | 14445 | 6882 | 47.6 |
C1 | No.4 (Shikoku) | No.5 (Tohoku) | 11062 | 1644 | 14.9 |
No.10 (Tohoku) | No.22 (Tohoku) | 10348 | 4981 | 48.1 |
No.22 (Tohoku) | No.44 (Kyushu) | 24017 | 4014 | 16.7 |
No.33 (Tohoku) | No.43 (Kyushu) | 34734 | 5058 | 14.6 |
No.43 (Kyushu) | No.4 (Shikoku) | 19758 | 2748 | 13.9 |
No.48 (Kanto) | No.4 (Shikoku) | 7961 | 4030 | 50.7 |
No.49 (Chubu) | No.4 (Shikoku) | 11033 | 2402 | 21.8 |
C2 | No.7 (Chubu) | No.11 (Chubu) | 13991 | 3218 | 23.0 |
No.7 (Chubu) | No.23 (Chubu) | 15078 | 3307 | 21.9 |
No.11 (Chubu) | No.23 (Chubu) | 15539 | 4533 | 29.2 |
No.11 (Chubu) | No.24 (Tohoku) | 13055 | 7026 | 53.8 |
No.23 (Chubu) | No.24 (Tohoku) | 16974 | 4967 | 29.3 |
No.24 (Tohoku) | No.7 (Chubu) | 15418 | 3432 | 22.2 |
C3 | No.1 (Kinki) | No.3 (Shikoku) | 13815 | 5548 | 40.1 |
No.3 (Shikoku) | No.20 (Chugoku) | 20615 | 3629 | 17.6 |
No.16 (Kyushu) | No.20 (Chugoku) | 25155 | 5146 | 20.4 |
No.18 (Chubu) | No.41 (Kyushu) | 12343 | 2167 | 17.5 |
No.37 (Kyushu) | No.45 (Kyushu) | 11183 | 2933 | 26.2 |
No.45 (Kyushu) | No.1 (Kinki) | 13057 | 2782 | 21.3 |
No.46 (Kyushu) | No.16 (Kyushu) | 25504 | 6161 | 24.1 |
C4 | No.6 (Kinki) | No.13 (Kinki) | 24481 | 3843 | 15.7 |
No.13 (Kinki) | No.28 (Kyushu) | 26763 | 3199 | 11.9 |
No.17 (Okinawa) | No.32 (Okinawa) | 12418 | 5361 | 43.9 |
No.19 (Chugoku) | No.38 (Kyushu) | 20175 | 2568 | 12.7 |
No.29 (Shikoku) | No.47 (Kinki) | 3007 | 131 | 4.3 |
No.47 (Kinki) | No.6 (Kinki) | 2473 | 136 | 5.4 |
No.50 (Kyushu) | No.6 (Kinki) | 12063 | 2425 | 20.1 |
We calculated the ratio of heterozygous loci for each individual (Supplementary Table S4). Ten individuals exhibited a relatively low ratio of heterozygous loci, with a value below 0.2. All ten plants (5, 7, 17, 28, 29, 33, 38, 43, 47, and 50) belonged to group C.
Each of groups A and B propagated asexually, while group C propagated sexually
A. macrostemon reproduces asexually via bulbs, particularly with human intervention. We detected asexual reproduction in the plant by examining the conservation of heterozygous loci positions via pairwise alignments. The degrees of conservation for the technical replicates, plants 14 and 15, 30 and 31, and 17 and 32, were 49.0%, 47.6%, and 43.9%, respectively. De novo analysis of RAD-Seq data, which contains many missing values, is likely to result in 40–50% conservation between somatic clones. Consequently, we determined the presence or absence of asexual reproduction by setting the value of conservation at nearly 40% as the criterion. It should be noted that a similar value of conservation was also used as a criterion for clonal propagation in a study of Japanese pepper that used de novo analysis of RAD-Seq data28.
We investigated conservation among plants in each group, both within and between districts (Table 4, Supplementary Table S5). The conservation level in group A was high, with over half of the pairs showing above 40% (40.8–52.1%). Some pairs had conservation levels slightly below 40% (36.6–39.7%). However, given the somatic mutations (including loss of heterozygosity) that occur during asexual reproduction, it is likely that these plants with slightly lower conservation values also propagated from the same ancestor via asexual reproduction. The conservation level in group B was high, with all pairs of plants showing an average of more than 40% (40.7–49.7%) except for one pair which showed 39.6%. On the other hand, nearly all the pairs observed in group C showed a low level of conservation, except for six instances. Thus, groups A and B propagated from a single plant by asexual reproduction, whereas the members of group C propagated almost exclusively without asexual reproduction.
Morphological differences exist among ancestral populations A, B, and C1.
Out of the 47 individuals subjected to RAD-Seq, 22 were cultured and analyzed for morphological data using nine distinct measurements (Table 5). We subsequently conducted a Principal Component Analysis (PCA) on the 22 cultured individuals, using the acquired morphological data (Fig. 6a). Given that groups A, B, and C1 represent ancestral populations, a separate PCA was performed exclusively on the ten individuals from these groups (Fig. 6b). The first PCA that included all 22 individuals accounted for 41.28% of the total variance, with the second principal component contributing 25.8%. Conversely, in the PCA exclusive to the ten selected individuals, the first and second principal components accounted for 58.4% and 23.3% of the variance, respectively. Notably, the graphical representation in Fig. 6a did not facilitate either genetic or regional grouping, while Fig. 6b allowed for genetic grouping exclusively. These observations underscore the presence of morphological disparities among the ancestral populations A, B, and C1.
Table 5
Morphological features of Allium macrostemon collected in various parts of Japan.
No. | Isolate | Group | Number of leaves | Plant number of tillers | Leaf length | Leaf width | Leaf sheath diameter | Number of daughter bulbs | Total weight of bulbs | Maximum weight of bulbs | Lateral diameter of maximum bulb |
39 | WOF366 | A | 7.3 ± 1.3 | 2.5 ± 1.3 | 44.8 ± 0.2 | 5.1 ± 0.4 | 8.5 ± 0.0 | 16.7 ± 3.3 | 19.8 ± 4.0 | 3.9 ± 0.0 | 19.7 ± 0.2 |
26 | WOKW581 | A | 10.4 ± 0.8 | 2.0 ± 0.2 | 58.0 ± 4.3 | 6.9 ± 0.6 | 10.5 ± 0.5 | 15.6 ± 0.0 | 30.2 ± 0.6 | 7.7 ± 0.1 | 23.8 ± 0.0 |
42 | WOOY575 | A | 9.3 ± 0.1 | 3.4 ± 1.6 | 58.7 ± 0.2 | 7.8 ± 0.6 | 12.5 ± 0.6 | 16.9 ± 3.5 | 31.5 ± 1.7 | 6.4 ± 2.1 | 22.1 ± 2.0 |
2 | WOGU466 | B | 8.4 ± 0.0 | 4.0 ± 0.4 | 61.4 ± 0.9 | 7.9 ± 0.3 | 12.9 ± 0.1 | 6.0 ± 0.4 | 22.1 ± 0.9 | 11.4 ± 0.9 | 26.8 ± 0.4 |
12 | WOIB449 | B | 9.9 ± 0.9 | 4.1 ± 0.7 | 58.9 ± 3.4 | 9.6 ± 1.5 | 16.2 ± 2.0 | 5.6 ± 0.4 | 23.6 ± 3.4 | 10.6 ± 0.4 | 26.9 ± 0.2 |
14 | WOKO594 | B | 9.1 ± 0.7 | 3.0 ± 0.0 | 56.9 ± 1.9 | 8.2 ± 0.6 | 14.4 ± 0.1 | 5.0 ± 0.0 | 24.3 ± 2.3 | 13.3 ± 3.6 | 28.6 ± 2.3 |
27 | WOME413 | B | 9.4 ± 1.2 | 4.3 ± 1.3 | 56.0 ± 5.0 | 8.5 ± 1.3 | 11.3 ± 1.5 | 4.6 ± 2.8 | 12.2 ± 5.6 | 6.9 ± 1.3 | 22.6 ± 1.8 |
5 | WOA360 | C1 | 5.1 ± 0.3 | 1.5 ± 0.1 | 45.7 ± 3.1 | 6.2 ± 0.0 | 7.4 ± 0.2 | 3.4 ± 0.2 | 8.5 ± 0.2 | 4.9 ± 0.1 | 19.5 ± 0.7 |
4 | WOEH562 | C1 | 7.6 ± 1.2 | 1.9 ± 0.5 | 49.3 ± 3.5 | 8.0 ± 0.5 | 14.6 ± 2.3 | 4.7 ± 1.1 | 15.9 ± 0.3 | 7.9 ± 1.7 | 23.9 ± 1.2 |
9 | WONR546 | C1 | 8.5 ± 1.3 | 1.8 ± 0.6 | 49.4 ± 0.7 | 7.6 ± 0.6 | 10.3 ± 2.2 | 4.6 ± 0.8 | 17.1 ± 0.4 | 7.3 ± 1.0 | 24.2 ± 0.5 |
23 | WOIS532 | C2 | 9.4 ± 0.2 | 2.5 ± 0.3 | 62.3 ± 1.9 | 6.3 ± 0.4 | 12.0 ± 0.5 | 9.2 ± 1.4 | 44.6 ± 22.6 | 16.4 ± 8.6 | 31.4 ± 5.7 |
7 | WOTY540 | C2 | 9.4 ± 2.1 | 2.0 ± 0.2 | 38.5 ± 3.8 | 5.3 ± 0.5 | 9.9 ± 0.6 | 14.0 ± 1.8 | 27.2 ± 8.0 | 4.8 ± 0.7 | 21.3 ± 0.1 |
18 | WOFI554 | C3 | 9.2 ± 0.4 | 8.5 ± 0.7 | 54.5 ± 0.8 | 8.7 ± 0.4 | 13.9 ± 0.1 | 76.3 ± 17.7 | 70.3 ± 0.7 | 5.6 ± 1.1 | 21.5 ± 1.4 |
16 | WOKG195 | C3 | 7.5 ± 0.7 | 3.5 ± 0.1 | 45.3 ± 2.2 | 7.1 ± 0.2 | 13.6 ± 0.8 | 6.1 ± 0.7 | 33.6 ± 1.8 | 14.3 ± 0.6 | 29.6 ± 0.4 |
1 | WOKY536 | C3 | 8.0 ± 1.2 | 5.8 ± 0.0 | 52.2 ± 3.9 | 8.1 ± 0.2 | 10.0 ± 1.7 | 10.9 ± 0.7 | 24.4 ± 1.8 | 5.8 ± 0.2 | 22.0 ± 0.4 |
20 | WOOY574 | C3 | 12.0 ± 2.4 | 5.5 ± 0.9 | 51.8 ± 4.5 | 7.9 ± 0.1 | 15.6 ± 1.5 | 13.2 ± 0.2 | 30.1 ± 14.9 | 5.4 ± 1.9 | 20.9 ± 1.9 |
40 | WOF395 | C4 | 9.1 ± 2.5 | 5.4 ± 2.8 | 53.7 ± 3.4 | 8.5 ± 0.3 | 14.9 ± 1.6 | 18.7 ± 11.3 | 45.9 ± 1.9 | 8.4 ± 2.4 | 24.9 ± 1.8 |
38 | WOM366 | C4 | 8.2 ± 0.6 | 3.3 ± 0.7 | 51.0 ± 0.9 | 7.5 ± 0.4 | 13.3 ± 1.9 | 20.8 ± 0.2 | 50.3 ± 17.1 | 7.3 ± 2.5 | 22.8 ± 2.5 |
28 | WOO199 | C4 | 9.7 ± 2.3 | 6.1 ± 0.7 | 69.6 ± 6.4 | 6.8 ± 0.8 | 13.9 ± 1.5 | 13.6 ± 0.8 | 44.7 ± 4.7 | 8.7 ± 1.0 | 25.2 ± 0.5 |
17 | WOOK122 | C4 | 7.2 ± 0.6 | 2.3 ± 0.3 | 53.5 ± 2.7 | 6.3 ± 0.2 | 11.0 ± 0.4 | 14.7 ± 0.3 | 32.6 ± 0.7 | 7.1 ± 0.7 | 25.0 ± 2.2 |
6 | WOSG538 | C4 | 10.7 ± 1.5 | 6.2 ± 0.2 | 76.0 ± 8.7 | 8.1 ± 0.8 | 10.6 ± 1.8 | 27.0 ± 8.0 | 57.6 ± 0.8 | 9.7 ± 2.2 | 25.0 ± 1.0 |
19 | WOT105 | C4 | 11.2 ± 1.2 | 4.4 ± 0.2 | 61.5 ± 1.1 | 7.4 ± 0.2 | 13.9 ± 0.4 | 16.7 ± 1.1 | 61.3 ± 1.2 | 9.4 ± 0.1 | 27.0 ± 1.5 |
Based on the results of the PCA, we further investigated which morphological measurements primarily influenced the groupings observed in Fig. 6b. Among the variables, leaf width, maximum weight of bulbs, lateral diameter of the maximum bulb, leaf sheath diameter, and the number of plant tillers were characterized by elevated values in group B. Conversely, the number of daughter bulbs demonstrated higher values within group A. Leaf length, the number of leaves, and the total weight of bulbs displayed enhanced values in both group A and group B, indicating shared morphological features among these two groups. Remarkably, none of the measurements presented high values for group C1, suggesting distinct morphological characteristics within this group.
We examined the preceding study33 that analyzed the concentration of phenolics in A. macrostemon collected from various regions across Japan. Using data extracted from this study33, we conducted a PCA, employing the quantitative values of functional components as variables. However, similar to the results in Fig. 6a, this PCA did not facilitate the delineation of samples based on genetic or regional clusters.