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Article

Population Structure and Genetic Diversity of Agave Germplasms in China

by
Xiaoli Hu
1,2,†,
Yubo Li
1,3,†,
Shibei Tan
1,†,
Lisha Chen
4,
Dietram Samson Mkapa
1,2,5,
Chen Lin
6,
Qingqing Liu
6,
Gang Jin
7,
Tao Chen
7,
Xu Qin
7,
Kexian Yi
8,* and
Xing Huang
1,9,10,*
1
National Key Laboratory for Tropical Crop Breeding, Environment and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, China
2
College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
3
School of Tropical Agricultural and Forestry, Hainan University, Danzhou 571737, China
4
Quality Inspection and Testing Center for Sisal Products, Ministry of Agriculture and Rural Affairs, Zhanjiang 524000, China
5
Mlingano Centre, Tanzania Agricultural Research Institute (TARI), Tanga P.O. Box 5088, Tanzania
6
Shanghai Key Laboratory of Plant Functional Genomics and Resources, Shanghai Chenshan Botanical Garden, Shanghai 201602, China
7
Guangxi Subtropical Crops Research Institute, Nanning 530001, China
8
Sanya Research Institute, Chinese Academy of Tropical Agricultural Sciences, Sanya 572025, China
9
Key Laboratory of Integrated Pest Management on Tropical Crops, Ministry of Agriculture and Rural Affairs, Haikou 571101, China
10
Hainan Key Laboratory for Monitoring and Control of Tropical Agricultural Pests, Haikou 571101, China
*
Authors to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Agronomy 2025, 15(3), 722; https://doi.org/10.3390/agronomy15030722
Submission received: 18 February 2025 / Revised: 9 March 2025 / Accepted: 14 March 2025 / Published: 17 March 2025
(This article belongs to the Special Issue Germplasm Conservation and Genetic Improvement in Tropical and Subtropical Crops)
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Versions Notes

Abstract

:
Agave hybrid cultivar 11,648 has been planted for sisal fiber production in China since the 1960s. However, little is known about the population structure and genetic diversity of agave germplasms in China. Therefore, we developed a group of core SNP markers to evaluate the population structure and genetic diversity of 125 agave germplasms in China, including 20 cultivars, 14 breeding lines, and 89 transplanted resources from different areas. Cost-effective amplicon sequencing technology was used to identify genetic variants. The results grouped most cultivars and breeding lines together, which indicated that local agave breeding programs aimed to improve fiber and disease-resistance traits. These breeding programs have reduced genetic diversity, even with the gene flows from other Agave species. The neighbor-joining phylogenetic tree revealed the relationships between A. H11648 and its parents. The phylogenetic relationship between A. sisalana and A. amanuensis is doubtful, even if they are considered heterotypic synonyms. The 11 agave germplasms introduced from Mexico suggest the abundant diversity of agave germplasms in Mexico, which is also the source of agave germplasms in China. This study provides a sketch map for agave germplasms in China, which will benefit future studies related to population genetics and breeding works of agave.

1. Introduction

The Agave genus is a typical tropical plant that is widely used in the production of spirits, sisal fiber, food, forage, medicine, cosmetics, and so on [1,2]. The genus originates from the American continent, where the tropical climate leads to its diversity, containing approximately 200 Agave species [3]. The long life cycle and vegetative reproduction make it important to reveal the evolution and adaptation derived from the genetic diversity of the genus [4,5]. The isozyme and allozyme systems have long been used to evaluate genetic diversity within or between Agave species, including Agave fourcroydes, A. angustifolia, A. victoriae-reginae, A. lechuguilla, A. murpheyi, A. delamateri, A. cocui, and A. parryi [6,7,8,9,10,11]. The phylogenetic relationships of Agave species have been efficiently characterized by ITS (Internal Transcribed Spacer) and chloroplast sequences (regional sequences or complete genomes) [4,12]. They are also used to analyze genetic diversity within Agave species, such as A. kerchovei, A. salmiana, A. mapisaga, and A. shawii [13,14,15]. Random amplified polymorphic DNA (RAPD) has revealed the genetic differentiation in the A. deserti complex, comprising A. deserti, A. cerulata, and A. subsimplex [16]. Inter-simple sequence repeat (ISSR) analysis has provided new insights into the conservation of A. angustifolia and the climate-derived differentiation of A. striata [17,18]. Amplified fragment length polymorphism (AFLP) analysis is an efficient tool for identifying agave genotypes but does not directly correlate with the disease resistance of Fusarium solani or Phytophthora nicotianae [19,20]. Microsatellites (simple sequence repeats, SSR) have become the major kind of molecular marker to estimate the genetic variance of Agave species during the past decade and have been successfully used in A. parryi, A. palmeri, A. utahensis, A. inaequidens, A. cupreata, A. hookeri, A. maximiliana, A. salmiana, A. mapisaga, A. americana, A. mapisaga, and five intraspecific entities [21,22,23,24,25,26]. In recent years, the rapid development of next-generation sequencing technology has become a powerful tool for genotyping, with a large number of markers at the genome level [27]. However, the large genomes of Agave species significantly increase the cost of genome assembly and resequencing [28]. Restriction site-associated DNA sequencing (RAD-Seq) is relatively economical and efficient for single nucleotide polymorphism (SNP) discovery and genotyping and has been used to reveal the genomic diversity in populations of A. tequilana, A. angustifolia, A. potatorum, and A. aurea [29,30,31,32,33]. To date, most studies related to genetic diversity have focused on agave populations around North America, the origin center of the genus [33]. Few similar studies have been reported from other continents, even though the hybrid cultivar A. H11648 ((A. amaniensis × A. angustifolia) × A. amaniensis) has been widely cultivated for sisal fiber production in Africa and Asia [34].
The hybrid cultivar A. H11648 was introduced in Tanzania in 1963 and has been cultivated over approximately 15 thousand hectares, with a dry fiber yield of 4.57 tons per hectare in 2021 [35]. This cultivar has long been vegetatively propagated in China, leading to a decline in yield and disease resistance. Thus, most agave-related studies have focused on fiber and disease-resistance traits in China [36]. A series of fiber-related genes have been characterized, including Cellulose Synthase, Small Auxin Up-regulated RNA, Cinnamyl Alcohol Dehydrogenase, Phenylalanine Ammonia-Lyase, and glycosyltransferase genes [1,28,34,37,38]. Chinese breeders have also selected a series of cultivars that are resistant to P. nicotianae, including the 76416, Guifu 4, Dong series, Nanya series, and Yuexi series [19]. In addition, several disease-resistance genes have been genetically transformed into A. H11648 to generate disease-resistant germplasms [39,40]. The genetic improvement aimed at fiber and disease resistance in China is completely different from that of A. tequilana in Mexico, which focuses on fructan-related traits [36]. However, there is no wild distribution of Agave species in China, which narrows its genetic diversity and restricts breeding procedures [35]. To date, little is known about the population structure and genetic diversity of the Agave germplasms in China. Hence, we collected 125 agave germplasms to evaluate their genetic backgrounds using cost-effective SNP-based amplicon sequencing [41]. The results will provide an overview of the population structure and genetic diversity of the agave germplasms in China, which will benefit future studies related to agave population genetics and breeding.

2. Materials and Methods

2.1. Sample Collection and DNA Isolation

Seedlings were collected from 125 agave germplasms, which were planted at the Environment and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences (Haikou, China, 19.99° N, 110.33° E). Among these, 18 and 83 were collected from the germplasm gardens of the Guangxi Subtropical Crops Research Institute (Nanning, China, 22.90° N, 108.35° E) and Quality Inspection and Testing Center for Sisal Products, Ministry of Agriculture and Rural Affairs (Zhanjiang, China, 21.40° N, 110.15° E), respectively (Table S1). There were 23 germplasms collected from Shanghai Chenshan Botanical Garden (Shanghai, China, 31.08° N, 121.19° E). The last germplasm (S102) was collected from the Environment and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences. We further classified these germplasms into three types according to their uses and origins, including 20 cultivars, 14 breeding lines, and 89 transplanted resources. The cultivars were created by artificial selection; they had been tested in field trials and could be cultivated for economic uses. The breeding lines were obtained by hybridization or mutagenesis and could be selected as cultivars after field trials. The transplanted resources were introduced from their original locations, which could be used as parent lines for hybridization. Notably, 11 transplanted resources were introduced from Mexico, including S16, S26, S43, S49, S50, S53, S57, S60, S69, S73, and S76. Leaf samples were collected from each germplasm, broken down into powder with liquid nitrogen, and further used for DNA isolation using the modified CTAB method [42]. Agarose gel electrophoresis was used to examine the quality of DNA samples to avoid degradation and RNA contamination. Nanodrop (Thermo Fisher Scientific, Waltham, MA, USA) was used to examine the purity of DNA samples with a cut-off for OD260/280 values between 1.8 and 2.2. Qubit 3.0 (Thermo Fisher Scientific, Waltham, MA, USA) was used to detect DNA concentrations (>20 ng/μL) and total amounts (>2 ug). The qualified DNA samples were stored at −80 °C until amplicon sequencing.

2.2. Marker Development and Amplicon Sequencing Analysis

A total of 100 SNP markers were selected according to the A. tequilana genome and resequencing data (unpublished), which were uniformly distributed on 30 chromosomes with interval sequences of over 20 Mb. Primers were designed using Primer 3 (version 2.5.0) for each marker, with primer lengths between 17 and 32 bp, Tm values between 60 and 64 °C, and amplicon sizes within 500 bp (Table S2) [43]. The KAPA2G Fast Multiplex Mix Kit (KAPA Biosystems, Cape Town, South Africa) was used for multiplex PCR, with mixed primers for each DNA sample, according to the manufacturer’s instructions. The annealing temperature was 62 °C with an extension time of 90 s and cycle numbers of 38. Adaptors were attached to amplicons, which were pooled and purified using AMPure XP Beads (Beckman Coulter, Brea, CA, USA) for library construction. The library was primarily quantified with Qubit 3.0, checked for insert sizes with Agilent 2100 (Agilent Technologies, Santa Clara, CA, USA), and accurately quantified with Analytik Jena qTOWER (Analytik Jena, Jena, Germany) to obtain a qualified library (>2 nM). The Illumina HiSeq 2500 platform (Illumina, San Diego, CA, USA) was selected for amplicon sequencing with paired-end 150 bp raw reads. The raw data were filtered using Fastp software (version 0.23.0) to remove adaptors and low-quality reads. The clean reads were mapped to reference sequences (Table S2) and aligned using Burrow-Wheeler Aligner with the MEM algorithm (version 0.7.15-r1140) [44,45]. The aligned reads were listed in SAM format files, which were transformed into BAM format files using SAMtools (version 1.3.1) [46]. The BAM files were used for variant calling, including SNP and InDel, using the GenotypeGVCFs module in the Genome Analysis Tool Kit (version 3.7) [47]. The variants were filtered with sequencing depth ≥ 10, percentage of genotype deletions ≤ 50%, and minor allele frequency ≥ 5%.

2.3. Population Structure and Genetic Diversity Analysis

STRUCTURE software (version 2.3.4) was used to analyze the population structure according to the genotypes from SNP calling [48]. ΔK values were calculated to estimate the subgroup numbers of the population when K values ranged from 1 to 7. The SNP variants were used to calculate genetic distances using MEGA7 (version 7.0), which was further used for principal component analysis (PCA) using PLINK (version v1.90p) [49,50]. A neighbor-joining phylogenetic tree was constructed according to genetic distances with MEGA7 (version 7.0) and visualized using Ggtree (version 1.7.10) [50,51]. VCFtools (version 0.1.17) was used to calculate indicators related to genetic diversity, including expected heterozygosity (He), polymorphism information content (PIC), and nucleotide diversity (pi), together with population differentiation (Fst) [52]. The He value ranged from 0 to 1, where 0 indicated the complete absence of polymorphism, meaning that the allele frequencies of all individuals in the population were the same; 1 indicated that there were an infinite number of alleles in the population, and the probability of each allele appearing was equal, which was the idealized maximum heterozygosity value. The ranges of the PIC and pi values were both between 0 and 1, where 0 indicated that all individuals had the same genotype at that locus and 1 indicated that each individual’s genotype was unique. The range of FST was [0, 1]. FST = 1 indicated that the allele was fixed in each subgroup and fully differentiated. FST = 0 meant that the genetic structure of different subgroups was completely consistent and there was no differentiation between populations.

3. Results

3.1. Genetic Variants by Amplicon Sequencing

A total of 100 pairs of primers were selected for the amplicon sequencing of agave germplasms from China (Table S1). The results indicated 122 loci with variants, including seven InDel and 114 SNP loci (Table S2). Among these, there was one nucleotide at the length of the seven InDel loci. The most abundant SNP types were A/G and C/T with counts of 42 and 37, respectively (Figure 1). In addition, the numbers of the other four types of SNP loci were within a dozen. There were 79 SNPs subjected to transitions (Ts), including A/G and C/T. The other four SNP types (35) were designated as transversions (Tv). The ratio of Ts/Tv was 2.26.

3.2. Population Structure of Agave Germplasms in China

We collected 125 agave germplasms to evaluate their genetic background, including 20 cultivars, 89 transplanted resources, and 14 breeding lines (Table S1). Among these, 83 and 18 were collected from Zhanjiang and Nanning, respectively, which also contained most cultivars and breeding materials with the aim of improving fiber and disease-resistance traits. In addition, 23 germplasms were collected from Shanghai, most of which were Agave species for ornamental use. The remaining germplasm was collected in Haikou. Notably, 11 transplanted resources were introduced from Mexico, including S16, S26, S43, S49, S50, S53, S57, S60, S69, S73, and S76.
We further calculated the ΔK values to obtain the most suitable number for subgroups, which indicated K = 2 as the most suitable value (Figure 2). The population structure was analyzed according to the K values (Figure 3 and Table S3). When K = 2, all germplasms were divided into two subgroups (G1 and G2). When K = 3, some of the germplasms in G1 were divided into the new subgroups G3, together with five germplasms from G2. When K = 4, 11 germplasms from the remaining G1 group were divided into the new subgroup G4.

3.3. Genetic Relationships of Agave Germplasms in China

PCA analysis was performed to confirm the results of the population structure, which clearly divided the subgroups G1 and G2 of the population structure (Figure 4, Table S4). PC1 values indicated variance between the two subgroups, together with the existence of several intermediate germplasms. PC2 values revealed differentiation among the germplasms, especially among those species with PC2 > 0.2, including A. univittata (C3 and C53), A. funkiana (C27), and A. garciae-mendozae (C36). A neighbor-joining phylogenetic tree was constructed based on genetic distances, which indicated three major clusters (Figure 5). G1 and G2 contained five germplasms and one germplasm, respectively. Besides, five subclusters with species-specific features were identified, including A. americana (I), A. fourcroydes (II), A. sisalana (III), A. angustifolia (IV), and A. H11648 (V). Most cultivars and breeding lines were grouped in G2 with similar PC1 values ranging from 0 to 0.1. The 11 transplanted resources from Mexico were distributed in each subgroup.

3.4. Genetic Diversity and Differentiation of Subgroups

Three genetic indices were calculated for each locus to evaluate the genetic diversity of the subgroups: He, PIC, and pi (Table S5). The He values ranged from 0 to 0.499 for the G1 subgroup and 0 to 0.5 for the G2 subgroup. The average He values were 0.329 and 0.158 for G1 and G2, respectively. The PIC values ranged from 0 to 0.374 and 0 to 0.375 for the two subgroups. The average PIC values were 0.265 and 0.134, respectively. The pi values of G1 ranged from 0 to 0.505 compared to 0 to 0.504 for G2. The average pi value of G1 was 0.332, which was higher than that of G2 (0.160). FST values were calculated to evaluate the differentiation of subgroups, which ranged from 0 to 0.862 with an average value of 0.332 (Table S5).

4. Discussion

4.1. Cost-Effective Characterization of Agave Germplasms

Molecular markers, including RFLP, RAPD, AFLP, ISSR, SSR, and SNP, are useful tools for population genetics [53]. The development of next-generation sequencing technology has brought about a revolution in the discovery and application of molecular markers, which has significantly increased the throughput and data size [27]. The application of molecular markers in agave follows the trend well, effectively revealing low genetic differentiation and heterozygote excess in Agave species [30,31]. However, most studies related to molecular markers have focused on the evolutionary patterns of agave populations, even with the precise application of marker-assisted selection in plant breeding [54]. Several scientists have tried but failed to find connections between AFLP markers and disease resistance in agave, which may be revealed by the high density of SNP markers in future studies [19,20]. It is also important to map quantitative trait loci (QTL) for fiber and fructan traits in agaves, which will facilitate breeding programs for this perennial tropical crop [36]. In addition, molecular markers are efficient tools in Distinctness, Uniformity, and Stability (DUS) systems, which are commonly used to characterize and protect plant varieties [55]. To date, few studies have reported the DUS system in agave. Therefore, we developed a group of core SNP markers to characterize agave germplasms in China. The results proved their effectiveness in identifying inter- and intraspecific features of agave germplasms (Figure 3 and Figure 4), which could be used as a guide for the DUS system in agave. However, the population structure results indicate that K = 2 is the best partition, which is not consistent with the three major branches in the phylogenetic tree. This could be due to the density of the SNP markers, which requires future studies. Moreover, amplicon sequencing is more cost-effective than resequencing and RAD-Seq, which will be a preferred tool for evaluating the genetic diversity of agave populations with large numbers of individuals [41].

4.2. Agave Breeding in China

The hybrid A. H11648 has been planted in Guangxi, Guangdong, and Hainan for over 60 years and is mainly used for sisal fiber production in China [35]. Thus, it is important to improve the fiber traits. Most cultivars and breeding lines were grouped together in the G2 population, which indicates the fiber-related breeding aim of agave in China (Figure 3). The results also revealed a narrow genetic background, which is closely related to A. H11648 and indicates its centered role in agave breeding in China (V in Figure 5). The identification of fiber-related genes has provided candidate targets to improve fiber traits, such as Cellulose Synthase, Cinnamyl Alcohol Dehydrogenase, Phenylalanine Ammonia-Lyase, and Glycosyltransferase genes [38]. Indeed, the cultivar is used to face the devasting pathogen P. nicotianae in China, which has promoted breeding programs for disease-resistant cultivars, including 76416, Guifu 4, Dong series, Nanya series, and Yuexi series [19]. The cultivar Nanya 2 (S65) suggests genetic introgression from A. americana (I in Figure 5), which is immune to P. nicotianae [19]. The cultivar Dong 16 (S82, S99) indicates a hybrid genetic background from A. karwinskii (C46). Several resistance genes have also been successfully transformed into A. H11648 to generate disease-resistant germplasms [39,40]. Furthermore, the results revealed the phylogenetic position of A. angustifolia, a parent of A. H11648 (IV in Figure 5). In contrast, the other parent, A. amanuensis, has a relatively complicated phylogenetic position, which might be caused by backcross and requires a larger density of SNP markers to locate it (V in Figure 5). Notably, A. sisalana is outside the subgroup of A. amaniensis, even though it is considered a heterotypic synonym (III in Figure 5). Their morphological traits are so similar that genomic proofs are still needed to reveal their relationships. Moreover, the 11 transplanted resources (S16, S26, S43, S49, S50, S53, S57, S60, S69, S73, and S76) introduced from Mexico were distributed in each subgroup, suggesting the abundant diversity of agave germplasms in Mexico and the source of agave germplasms in China (Figure 5). We further evaluated their genetic diversity and differentiation, which indicated decreased He, PIC, and pi indicators in the G2 group of cultivars and breeding lines (Table S5). This is closely related to A. H11648-centered genetic improvement in China. The FST value indicated relatively low differentiation between the G1 and G2 subgroups, which indicated strong gene flow from the G1 subgroup [56]. This is consistent with the disease resistance introgression from A. americana and similar to the genetic differentiation of A. angustifolia populations (0.3747 and 0.24) according to previous studies [29,31]. In contrast, population genomics suggests extremely low genetic differentiation in A. tequilana (0.0044), A. potatorum (0.0796), and A. aurea (0.087) [30,32,33]. The findings indicate that interspecific crosses might be an efficient way to increase genetic diversity for agave species, which emphasizes the importance of protecting the diversity of agave species. This study provides a sketch map for agave germplasms in China, which will benefit future agave breeding studies.

5. Conclusions

In this study, we developed a group of core SNP markers to evaluate the population structure and genetic diversity of 125 agave germplasms from China. The results grouped most cultivars and breeding lines together, which indicated local agave breeding programs aimed at improving fiber and disease-resistance traits. These breeding programs have reduced genetic diversity even with gene flow from other Agave species. The neighbor-joining phylogenetic tree revealed the relationships between A. H11648 and its parents. There remains doubt about the phylogenetic relationship between A. sisalana and A. amaniensis, even if they are considered heterotypic synonyms. The eleven agave germplasms introduced from Mexico suggest the abundant diversity of agave germplasms in Mexico, which is also the source of the agave germplasms in China. This study provides a sketch map for agave germplasms in China, which will benefit future studies related to population genetics and breeding works in agave.

Supplementary Materials

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/agronomy15030722/s1: Table S1: Agave germplasms used in this study, including 20 cultivars, 14 breeding lines, and 89 transplanted resources; Table S2: Primers and reference sequences covering 100 SNP loci; Table S3: Structure analysis of agave germplasms with K = 2, 3, and 4, respectively; Table S4: PCA analysis of agave germplasms. PC1 values indicate the variance between subgroups. PC2 values reveal the differentiation among germplasms; Table S5: Genetic diversity (He, PIC, and pi) of G1 and G2 subgroups and genetic differentiation (FST) between G1 and G2 subgroups.

Author Contributions

Conceptualization, X.H. (Xiaoli Hu), Y.L., S.T., L.C., D.S.M., C.L., Q.L., G.J., T.C., X.Q. and X.H. (Xing Huang); formal analysis, X.H. (Xiaoli Hu), Y.L., S.T., D.S.M., L.C., C.L., Q.L., G.J., T.C., X.Q., and X.H. (Xing Huang); investigation, X.H. (Xiaoli Hu), Y.L., S.T., L.C., C.L., Q.L., G.J., T.C., X.Q. and X.H. (Xing Huang); writing—original draft preparation, X.H. (Xiaoli Hu), Y.L., S.T. and X.H. (Xing Huang).; writing—review and editing, D.S.M., K.Y. and X.H. (Xing Huang); supervision, K.Y. and X.H. (Xing Huang); funding acquisition, K.Y. and X.H. (Xing Huang). All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Hainan Provincial Natural Science Foundation of China (323RC544, 322MS112), National Natural Science Foundation of China (32001598), China Agriculture Research System of MOF and MARA (CARS-16), and Central Public-interest Scientific Institution Basal Research Fund (1630042022005).

Data Availability Statement

All data are contained within the article or Supplementary Materials.

Acknowledgments

We would like to thank Bo Wang from Genoseq Technology Co., Ltd. (Wuhan, China) for his thorough suggestions for the experiment design and technical support.

Conflicts of Interest

The authors declare no conflicts of interest.

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Agronomy 15 00722 g001
Figure 1. SNP types and their counts by amplicon sequencing of Agave genus.
Figure 1. SNP types and their counts by amplicon sequencing of Agave genus.
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Figure 2. ΔK values with change in subgroup number (K).
Figure 2. ΔK values with change in subgroup number (K).
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Figure 3. Population structure for K = 2, 3, and 4. When K = 2, all germplasms were divided into two subgroups (G1 and G2). When K = 3, a part of the germplasms in G1 was divided into the new subgroup G3, together with five germplasms from G2. When K = 4, eleven germplasms from the rest of G1 were divided into the new subgroup G4.
Figure 3. Population structure for K = 2, 3, and 4. When K = 2, all germplasms were divided into two subgroups (G1 and G2). When K = 3, a part of the germplasms in G1 was divided into the new subgroup G3, together with five germplasms from G2. When K = 4, eleven germplasms from the rest of G1 were divided into the new subgroup G4.
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Figure 4. PCA analysis of genetic distance. The subgroups were labeled G1 and G2 according to population structure. PC1 values indicated the variance between subgroups. PC2 values revealed the differentiation among germplasms.
Figure 4. PCA analysis of genetic distance. The subgroups were labeled G1 and G2 according to population structure. PC1 values indicated the variance between subgroups. PC2 values revealed the differentiation among germplasms.
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Figure 5. Neighbor-joining phylogenetic tree based on genetic distances. The subgroups were labeled with G1 and G2 according to population structure. Five subclusters with species-specific features were identified, including A. americana (I), A. fourcroydes (II), A. sisalana (III), A. angustifolia (IV), and A. H11648 (V).
Figure 5. Neighbor-joining phylogenetic tree based on genetic distances. The subgroups were labeled with G1 and G2 according to population structure. Five subclusters with species-specific features were identified, including A. americana (I), A. fourcroydes (II), A. sisalana (III), A. angustifolia (IV), and A. H11648 (V).
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MDPI and ACS Style

Hu, X.; Li, Y.; Tan, S.; Chen, L.; Mkapa, D.S.; Lin, C.; Liu, Q.; Jin, G.; Chen, T.; Qin, X.; et al. Population Structure and Genetic Diversity of Agave Germplasms in China. Agronomy 2025, 15, 722. https://doi.org/10.3390/agronomy15030722

AMA Style

Hu X, Li Y, Tan S, Chen L, Mkapa DS, Lin C, Liu Q, Jin G, Chen T, Qin X, et al. Population Structure and Genetic Diversity of Agave Germplasms in China. Agronomy. 2025; 15(3):722. https://doi.org/10.3390/agronomy15030722

Chicago/Turabian Style

Hu, Xiaoli, Yubo Li, Shibei Tan, Lisha Chen, Dietram Samson Mkapa, Chen Lin, Qingqing Liu, Gang Jin, Tao Chen, Xu Qin, and et al. 2025. "Population Structure and Genetic Diversity of Agave Germplasms in China" Agronomy 15, no. 3: 722. https://doi.org/10.3390/agronomy15030722

APA Style

Hu, X., Li, Y., Tan, S., Chen, L., Mkapa, D. S., Lin, C., Liu, Q., Jin, G., Chen, T., Qin, X., Yi, K., & Huang, X. (2025). Population Structure and Genetic Diversity of Agave Germplasms in China. Agronomy, 15(3), 722. https://doi.org/10.3390/agronomy15030722

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