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Article

Genetic Diversity of Carpinus tientaiensis Cheng, an Endemic and Critically Endangered Species in China, Based on ITS Sequences

by
Runan Zhao
1,2,
Qianqian He
3,
Xiaojie Chu
4,
Anguo He
5 and
Zunling Zhu
1,2,6,*
1
College of Landscape Architecture, Nanjing Forestry University, Nanjing 210037, China
2
Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing 210037, China
3
School of Architecture, Zhijiang College of Zhejiang University of Technology, Shaoxing 312030, China
4
College of Life Sciences, Zhejiang Normal University, Jinhua 321004, China
5
Administration of Zhejiang Dapanshan National Nature Reserve, Jinhua 322300, China
6
Jinpu Research Institute, Nanjing Forestry University, Nanjing 210037, China
*
Author to whom correspondence should be addressed.
Forests 2023, 14(8), 1600; https://doi.org/10.3390/f14081600
Submission received: 29 June 2023 / Revised: 30 July 2023 / Accepted: 4 August 2023 / Published: 8 August 2023
(This article belongs to the Section Genetics and Molecular Biology)
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Abstract

:
The habitat of Carpinus tientaiensis Cheng (Betulaceae), an endemic endangered species in China, has been severely damaged, and it is in danger of going extinct. It is of great practical significance to propose corresponding protection measures based on population genetic variation. Based on the nuclear internal transcribed spacer (ITS) sequences, this study discovered that C. tientaiensis has relatively high genetic diversity at the species level. At the population level, the genetic variation levels of each population were not consistent, and the genetic diversity of the northern populations was higher than that of the southern populations. There was no significant genetic differentiation and phylogeographic structure among ribotypes and populations. Phylogenetic analysis showed that Sect. Distegocarpus and Sect. Carpinus were two independent genetic groups among the Carpinus, and C. tientaiensis may be the more evolutionary species in the Sect. Carpinus. Therefore, this study inferred that while C. tientaiensis has certain adaptability, climate change and human interference have brought it to an endangered state. Its populations may experienced the bottleneck effect, after which the expansion time was too short, with the populations failing to form a complex genetic structure. In addition, Tiantai Mountain was probably the original community and center of C. tientaiensis.

1. Introduction

Genetic diversity can represent the heritable variation within and between populations of organisms, including plants, and is essential for the conservation, utilization, selection, and improvement of plants [1,2]. When species have high genetic diversity, their adaptability to the environment is stronger, and the lower the diversity, the weaker the adaptability of species to the environment [3,4]. To a certain extent, research on genetic diversity can analyze the origins of species and speculate on the ability and mechanisms of species evolution [3,5]. Therefore, conducting genetic diversity research on endangered or rare species can not only elucidate the origin and evolutionary mechanisms of endangered species, but also the mechanisms by which they become endangered, and provide a theoretical basis for their protection and genetic diversity preservation [6,7].
Genetic diversity has emerged as one of the primary research topic in conservation biology because it is a key component of biodiversity, and biodiversity protection is ultimately about protecting genetic diversity [3]. The internal transcribed spacer (ITS) sequence of ribosomal DNA (rDNA) has the advantages of biparental inheritance, small fragment and conservative length, high copy number, fast base variation rate, and strong primer versatility [8]. The China Plant BOL (Barcode of Life) Group, based on the research of the CBOL (Consortium for the Barcode of Life) Plant Working Group, having evaluated the universality of DNA barcoding in 6286 individuals of 1757 species in 141 genera of 75 families of seed plants, found that the ITS sequence has high universality in angiosperms, and suggested that it be included in the core barcode for seed plants [9]. Nowadays, the ITS sequence is widely used to study species’ genetic diversity, genetic differentiation, and phylogenetic relationships at the inter-genus, inter-species, and even below-species levels [10,11], thus playing an important role in phylogenetic relationships and evolution of plants, as well as in conservation biology [12].
Carpinus tientaiensis Cheng, a deciduous tree of Betulaceae [13,14,15], is an endemic Tertiary endangered plant [14,15], which is only distributed in Zhejiang Province, China. It is listed on the IUCN (International Union for Conservation of Nature) Red List of Threatened Species as a critically endangered (CR) species [16]. It has important scientific and ecological value in the fields of phytogeography, species evolution, environmental adaptability of plants, and the formation and maintenance of biodiversity. However, its habitat is shrinking and there is a risk of extinction. It is of great practical significance to carry out genetic diversity research on C. tientaiensis for its resource protection and utilization. Therefore, this study aims to: (1) analyze the level of genetic variation and the population dynamics of C. tientaiensis based on ITS sequences; (2) preliminarily analyze its phylogenetic relationships; (3) propose strategies for the protection and management of germplasm resources.

2. Materials and Methods

2.1. Acquisition of Research Materials

A thorough field survey was conducted in Zhejiang and the adjacent areas between 2018 and 2020, covering all areas where C. tientaiensis may be distributed, and the longitude, latitude, and altitude of each population were recorded by handheld GPS. The study collected leaves from all adult individuals in each population, and the collected leaves were immediately stored in a preservation bag containing discolored silicone. In the end, leaf samples of C. tientaiensis were collected from Tiantai Mountain (TTS), Dapanshan (DPS), Yangtianhe (YTH), and Shangshantou (SST) (Figure 1; Table 1). The voucher specimens for each population were kept in the landscape experimental training center at Nanjing Forestry University (NFU), Nanjing, China.

2.2. Amplification and Sequencing

The modified CTAB method (modified from [17]) was used to extract the total DNA. PCR amplification was performed on all collected individuals, and the primer sequences [18] are shown in Table S1. The amplification system was 30 μL and contained 1 μL DNA, 27 μL Tsingke PCR Mix, 1 μL forward and reverse primers with a concentration of 2 μmol/L, and 1 μL template DNA with a concentration of 20 ng/μL. The reaction procedure of amplification was as follows: pre-denaturation at 98 °C for 3 min, followed by amplification for 38 cycles. The reaction system for each cycle was as follows: denaturation at 98 °C for 10 s, followed by primer annealing at 56 °C for 45 s, and then extension at 72 °C for 10 s, and extension at 72 °C for 7 min after all the cycles were completed. The PCR products were detected by electrophoresis, and then the products were sequenced on an ABI 3730 DNA sequencer (Applied Biosystems, Foster, CA, USA) by Tsingke Biotechnology Co., Ltd. (Beijing, China) (www.tsingke.com.cn, accessed on 28 September 2022), and, finally, 52 sequences were obtained (Table 1).

2.3. Analysis of Genetic Diversity

The sequencing results were manually corrected using the SeqMan Pro v11.1.0 software (DNASTAR Inc., Madison, WI, USA), and then all sequences were aligned using MEGA v10.1.7 software [19]. DNASP v5.10 software [20] was used to calculate the ribotype numbers, ribotype diversity (Hd), nucleotide polymorphisms (π), and genetic differentiation coefficients between populations (GammaSt), while gene flow (Nm) was estimated using the formula Nm = (1/GammaSt − 1)/2 [21]. POPART v1.7 software [22] was used to create the median-joining (MJ) network of ribotypes and analyze the relationships between ribotypes. Spatial interpolation analysis of population genetic diversity based on the Hd and IDW (inverse distance weighting) method [23] was performed using ArcGIS v10.2 software (ESRI Inc., Redlands, CA, USA).
PERMUT v2.0 software [24] was used to calculate the total genetic diversity (HT), the genetic differentiation coefficient between populations (GST and NST), and to compare their differences using 10,000 times replacement tests in order to ascertain whether the population had a significant phylogeographic structure. To determine if the population has experienced expansion, Tajima’s D test, Fu’s Fs test, and mismatch distribution analysis (MDA) were performed using DNASP v5.10 software [20].

2.4. Phylogenetic Analysis

In order to study the phylogenetic relationships of C. tientaiensis, this study also collected other species of Carpinus, which are distributed in the same or adjacent areas as C. tientaiensis. Due to some individuals sequencing failures, or only 1–2 individuals in certain populations of certain species being successfully sequenced, while populations with less than 3 sequences will be excluded, resulting in a total of 40 sequences being obtained (Table 1). DNASP v5.10 software was used to identify the ribotypes, and then POPART v1.7 software was used to create the MJ network. In addition, for phylogenetic analysis, some ITS sequences were obtained from NCBI (National Center for Biotechnology Information, https://www.ncbi.nlm.nih.gov/, accessed on 1 February 2023), with at least 2 sequences selected for each species, resulting in a total of 10 sequences obtained (Table 2). Phylogenetic analysis was performed using the NJ method (1000 bootstrap replications) in MEGA v10.1.7 software.

3. Results

3.1. Genetic Diversity and Ribotype Variation

In this study, a total of 52 ITS sequences of C. tientaiensis from four locations were obtained, which consisted of 679 bp. A total of nine ribotypes and eight polymorphic sites were detected, including five singleton variable sites and three parsimony informative sites (Table 3). At the species level, the total Hd of C. tientaiensis was 0.379 ± 0.086, and the total π was 0.720 × 10−3. At the population level, the range of Hd of each population was 0 to 0.507 ± 0.140, and the range of π was 0 to 0.840 × 10−3. Among them, the Hd (0.507 ± 0.140) and π (0.840 × 10−3) of the TTS population were the highest, while the YTH population had the lowest Hd and π, both of which were 0 (Table 4).
The genetic diversity of other Carpinus species collected through the field survey was tested. It was found that at the species level, the total Hd of C. tschonoskii was 0.500 ± 0.265, and the total π was 0.730 × 10−3. The total Hd of C. polyneura was 0.333 ± 0.215, and the total π was 0.490 × 10−3. The total Hd of C. hupeana was 0.533 ± 0.172, and the total π was 0.780 × 10−3. The total Hd of C. viminea was 0.221 ± 0.121, and the total π was 0.320 × 10−3. The total Hd and π of C. putoensis and C. cordata were 0. At the population level, the Hd (0.533 ± 0.172) and π (0.780 × 10−3) of the Shangshantou population of C. viminea (SSTL) were the highest, and the Hd and π of the other populations were all 0.

3.2. Ribotype Distribution and Network Structure

According to the MJ network of ribotypes of C. tientaiensis (Figure 2), there was no obvious genetic differentiation among ribotypes and populations. Among all ribotypes, R2 (41/52 = 78.85%) was the most common ribotype and widely distributed in all populations. It was speculated to be an ancestor ribotype. All other ribotypes had mutated from R2, where R3, R4, R5, and R6 were unique ribotypes of the SST population, and R7 and R9 were unique ribotypes of the TTS population.

3.3. Gene Flow and Genetic Structure

The gene flow (Nm) between different populations ranged from 4.81463 to 202.75203, with the SST and YTH populations having the most active gene flow, and the YTH and TTS populations having the least gene flow (Table S2; Figure 3). Spatial interpolation analysis revealed that the genetic diversity of the northern population was higher, while that of the southern population was lower (Figure 4). The total HT of C. tientaiensis was 0.309. The genetic differentiation coefficients between populations, GST and NST, were −0.001 and −0.016, respectively. Both GST and NST were negative values with no statistical significance (p = 0.0988) and NST values were lower than GST, indicating that there was no significant phylogeographic structure between populations.

3.4. Population Historical Dynamics

The result of the MDA of C. tientaiensis was a single peak (Figure 5), and the neutral test resulted in negative values with no statistical significance (Tajima’s D = −1.09883 (p > 0.1); Fu and Li’s D* = −1.85254 (p > 0.1)). Therefore, the populations of C. tientaiensis may not have experienced large-scale expansion recently.

3.5. Phylogenetic Analysis

In this study, a total of 92 ITS sequences of seven species were obtained, which consisted of 681 bp. Among these sequences, 19 ribotypes and 57 polymorphic sites were identified, including 7 singleton variable sites and 50 parsimony information sites (Table S3). Among them, C. tientaiensis has nine ribotypes (R1–R9), C. putoensis (R10) and C. cordata (R19) each have one ribotype, while C. tschonoskii (R11–R12), C. hupeana (R13–R14), C. polyneura (R15–R16), and C. viminea (R17–R18) all have two ribotypes (Table S3; Figure 6).
According to the MJ network (Figure 6), C. cordata and C. viminea were connected by 16 mutation steps and evolved into genetic branches independently. Therefore, the network supported the conclusion that Carpinus can be divided into Sect. Distegocarpus and Sect. Carpinus. C. viminea and mv1 (hypothetical haplotype 1) were connected by six mutation steps, and C. hupeana and C. polyneura were connected with mv3 by two mutation steps. It is speculated that C. hupeana and C. polyneura may have a common ancestor although they both evolved into genetic branches independently. In addition, a total of nine hypothetical haplotypes (mv1–mv9) were detected in six species of Sect. Carpinus.
Afterward, phylogenetic analysis was conducted using 19 ribotypes and 10 ITS sequences obtained from the NCBI, and the 29 sequences consisted of 617 bp. The phylogenetic results (Figure 7) supported the result of the MJ network (Figure 6). The Sect. Distegocarpus and Sect. Carpinus independently evolved into two genetic branches. The C. tientaiensis had a close genetic relationship with C. tschonoskii and C. putoensis, while C. hupeana and C. polyneura had a close genetic relationship.

4. Discussion

C. tientaiensis is a unique Tertiary endangered species in China, and its wild populations are only distributed in Zhejiang in eastern China [13,14,15]. The number of its wild individuals is very small, and the populations are spaced far apart, posing a risk of extinction. Previous study has found that the average value of nuclear gene variation in angiosperm was 0.137 [25]. Based on ITS sequences, it was found that the total genetic diversity (HT) of C. tientaiensis was 0.309, indicating that it had high genetic diversity at the species level. Some studies have shown that some endangered species have lower genetic diversity [26,27], while others have shown that some endangered species have higher genetic variation [28,29,30,31]. Researches have shown that the genetic diversity and genetic structure of species were affected by various factors such as distribution range, reproduction mode, evolutionary history, climate, and human interference [32,33,34,35], so endangered plants do not necessarily show low genetic diversity. Therefore, it is particularly necessary to identify the genetic diversity of the total and each population, and propose strategies for genetic diversity protection.
The nuclear genes were biparental inheritance [36], and extensive crossing led to genetic recombination, increasing the possibility of genetic variation. There is widespread natural hybridization and infiltration of anemophilous plants [37], and Betulaceae species are typical wind-pollinated plants, which have widespread hybridization and infiltration [38,39]. The same is true of the Carpinus species [39,40,41], which greatly promotes hybridization and gene exchange. Carpinus plants are Tertiary species, which have experienced a long evolutionary history. At the same time, C. tientaiensis is polyploid [42], which can accommodate and accumulate more variation [43], thus maintaining relatively high genetic diversity. However, study has found that polyploids have a higher mortality rate than diploids [44], and factors such as instability in polyploid genomes, abnormal mitosis and meiosis, and harmful effects of duplicate genes are not conducive to the survival of polyploids [45]. This reminded us of the need to strengthen the protection of polyploid species, especially those in endangered species, to study the ecological adaptability and maintenance mechanisms of polyploid species, and to provide scientific theories and methods for their protection.
The gene flow (Nm) among populations of C. tientaiensis was between 4.81463 to 202.75203, with a high gene flow. Frequent gene flow may lead to small or even no genetic differentiation or phylogeographic structure between populations [46,47]. The distribution range of C. tientaiensis is narrow, and the characteristics of wind pollination and samara spreading easily under the effect of wind have promoted pollination and seed transmission to a certain extent, resulting in a high gene flow. Therefore, there was no significant genetic differentiation and phylogeographic structure between the populations. The MJ network and the coefficient of genetic differentiation among populations (GST and NST) also confirm this conclusion. Research has shown that the ancient ribotype is generally located in the center of the ribotype network, with high distribution frequency and wide geographical distribution range [48], so it was speculated that R2 was the ancient ribotype. However, there was no significant genetic differentiation and phylogeographic structure among ribotypes, so it was hypothesized that the populations of C. tientaiensis experienced a bottleneck effect, after which the population expanded for a short time, failing to form a complex genetic structure. In addition, due to the existence of the “founder effect”, the genetic diversity of the original community was greater than that of the migration diffusion community, so the original community also had more genetic diversity and unique ribotypes [49]. Therefore, Tiantai Mountain may be the original community and center of C. tientaiensis. With the dynamic changes in climate, they gradually migrated toward the southwest and eventually spread to other regions.
It is generally believed that Carpinus can be divided into Sect. Distegocarpus and Sect. Carpinus [14,39,40,41,50], and this study confirmed this conclusion. It was found that C. viminea was a diploid species [51], however, C. tientaiensis was polyploid, which is the highest ploidy level in Betulaceae [42]. Based on this, it was assumed that among the species in Sect. Carpinus, C. viminea may be a relatively original species, and C. tientaiensis may be a relatively evolved species. In previous studies on the phylogeny of Carpinus based on ITS, it was found that C. tientaiensis has a close genetic relationship with C. viminea, while C. hupeana, C. tschonoskii, and C. putoensis have a close genetic relationship [52,53]. Recent research has found that C. tientaiensis has a close genetic relationship with C. tschonoskii, but a distant genetic relationship with C. viminea [54]. However, due to the limited number of species and sequences used in these studies, the genetic relationships of Carpinus still need further study. As more species were included in the phylogenetic analysis, the genetic relationships of Carpinus were further clarified. The study found that C. tientaiensis has a close genetic relationship with C. putoensis and C. tschonoskii, C. hupeana has a close genetic relationship with C. polyneura, and C. viminea has a distant genetic relationship with them [55]. These conclusions were consistent with the results of this research.
It is of great theoretical and practical significance to carry out genetic diversity research on C. tientaiensis for its protection and genetic diversity conservation. Tiantai Mountain is a famous scenic spot in China. Its tourism-related construction and sightseeing activities may have a serious impact on the survival of C. tientaiensis. Shangshantou is located in a remote and uninhabited area, currently with the largest wild population. Dapanshan is a national nature reserve that has not yet been developed for tourism, but individuals are spaced far apart. The Yangtianhe has built a reservoir, which is likely to damage the original community. This study found that there were differences in genetic diversity among populations, which also reminded us to focus on protecting wild individuals in Dapanshan and Yangtianhe. Therefore, scientific research should be continued to analyze the mechanisms of survival and environmental adaptability of C. tientaiensis, and strengthen its maintenance and management.

5. Conclusions

In this study, ITS sequences were used to evaluate the genetic diversity and phylogenetic relationships of C. tientaiensis. It was found that there was no obvious genetic differentiation and phylogeographic structure among ribotypes and populations, and R2 was the ancestral ribotype. The populations of C. tientaiensis may have experienced a bottleneck effect and failed to form a complex genetic structure, and may not have experienced large-scale expansion recently. There were differences in genetic diversity among populations, with higher genetic diversity in the northern populations and lower genetic diversity in the southern populations. Tiantai Mountain was probably the original community and center of C. tientaiensis.
At the nuclear gene level, C. tientaiensis had relatively high genetic variation, which indicated that C. tientaiensis may have strong environmental adaptability. However, due to human activities such as the construction of scenic spots, dams, and reservoirs, as well as windy and rainy environmental conditions, the fruiting and community regeneration of C. tientaiensis were affected, resulting in its endangered status. Therefore, it is necessary to strengthen the protection and management of C. tientaiensis, for example, to establish core reserves and carry out artificial breeding, so as to alleviate its habitat threat and endangered status.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/f14081600/s1, Table S1: Primer sequences and annealing temperature; Table S2: Coefficient of genetic differentiation (GammaSt) and gene flow (Nm) of C. tientaiensis; Table S3: Nineteen ribotypes and variable sites of seven Carpinus species.

Author Contributions

Conceptualization, R.Z.; methodology, R.Z.; formal analysis, R.Z., Q.H. and X.C.; investigation, R.Z., Q.H. and A.H.; data curation, R.Z.; writing—original draft preparation, R.Z.; writing—review and editing, R.Z., Q.H. and X.C.; supervision, Z.Z.; project administration, R.Z. and Z.Z.; funding acquisition, R.Z. and Z.Z. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Postgraduate Research and Practice Innovation Program of Jiangsu Province, grant number KYCX22_1104; the Forestry and Grassland Science and Technology Achievements National Promotion Project, grant number 2019[19]; the National Natural Science Foundation of China, grant number 31770752; and the “333” Scientific Research Project in Jiangsu Province, grant number BRA2018065.

Data Availability Statement

The 19 ribotypes sequences presented in this study have been uploaded to the National Center for Biotechnology Information [https://www.ncbi.nlm.nih.gov/GenBank ID: OR353711 to OR353729]. Publicly available datasets were analyzed in this study, and this data can be found here: [https://www.ncbi.nlm.nih.gov/GenBank ID: MG569958, MG727560, GQ250099, KC412170, AF297351, FJ011745, AY761128, FJ011778, AF098428, MN722083].

Conflicts of Interest

The authors declare no conflict of interest.

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Forests 14 01600 g001
Figure 1. Geographical locations of the sampling sites. JS: Jiangsu Province; AH: Anhui Province; ZJ: Zhejiang Province; NFU: Nanjing Forestry University, Nanjing City; TTZ: Tiantangzhai Scenic Spot, Lu’an City; PTS: Mount Putuo, Zhoushan City; TTS: Tiantai Mountain, Tiantai County; DPS: Dapan Mountain, Pan’an County; YTH: Yangtianhe, Qingtian County; SST: Shangshantou, Jingning She Autonomous County.
Figure 1. Geographical locations of the sampling sites. JS: Jiangsu Province; AH: Anhui Province; ZJ: Zhejiang Province; NFU: Nanjing Forestry University, Nanjing City; TTZ: Tiantangzhai Scenic Spot, Lu’an City; PTS: Mount Putuo, Zhoushan City; TTS: Tiantai Mountain, Tiantai County; DPS: Dapan Mountain, Pan’an County; YTH: Yangtianhe, Qingtian County; SST: Shangshantou, Jingning She Autonomous County.
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Figure 2. Median-joining (MJ) network of ribotypes of C. tientaiensis.
Figure 2. Median-joining (MJ) network of ribotypes of C. tientaiensis.
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Figure 3. GammaSt and Nm of C. tientaiensis. Above the diagonal is Nm, and below the diagonal is GammaSt.
Figure 3. GammaSt and Nm of C. tientaiensis. Above the diagonal is Nm, and below the diagonal is GammaSt.
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Figure 4. Spatial interpolation analysis of C. tientaiensis.
Figure 4. Spatial interpolation analysis of C. tientaiensis.
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Figure 5. The result of mismatch distribution analysis (MDA) of C. tientaiensis.
Figure 5. The result of mismatch distribution analysis (MDA) of C. tientaiensis.
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Figure 6. Median-joining (MJ) network based on 19 ribotypes.
Figure 6. Median-joining (MJ) network based on 19 ribotypes.
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Figure 7. Phylogenetic relationships of 19 ribotypes and 10 ITS sequences obtained from NCBI.
Figure 7. Phylogenetic relationships of 19 ribotypes and 10 ITS sequences obtained from NCBI.
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Table 1. Collection records and numbers of sequences.
Table 1. Collection records and numbers of sequences.
IdentifierPopulationsGeographic LocationLongitudeLatitudeAltitudeNumberSpecies
1-1–1-5DPSDapanshan National Nature Reserve, Pan’an, Zhejiang120.521828.970811385C. tientaiensis
2-1–2-17TTSTiantai Mountain, Tiantai County, Zhejiang121.091729.256890117C. tientaiensis
3-1–3-2YTHYangtianhe, Qingtian, Zhejiang119.990728.208412492C. tientaiensis
4-1–4-28SSTShangshantou, Jingning She Autonomous County, Zhejiang119.632027.7823150628C. tientaiensis
P1-1PTSPMount Putuo, Zhoushan, Zhejiang122.397730.01772921C. putoensis
P1-2–P1-4PTSPNanjing Forestry University, Nanjing, Jiangsu (introduced from Mount Putuo)118.827632.0852243C. putoensis
L1-1–L1-3DPSLDapanshan National Nature Reserve, Pan’an, Zhejiang120.522928.974711363C. viminea
L2-1–L2-6TTSLTiantai Mountain, Tiantai County, Zhejiang121.090829.25259266C. viminea
L3-1–L3-3YTHLYangtianhe, Qingtian, Zhejiang119.990728.208412493C. viminea
L4-1–L4-5SSTLShangshantou, Jingning She Autonomous County, Zhejiang119.630927.784015185C. viminea
H1-1–H1-6TTZHTiantangzhai Scenic Spot, Lu’an, Anhui115.762531.15677606C. hupeana
D1-1–D1-6DPSDDapanshan National Nature Reserve, Pan’an, Zhejiang120.522628.973210866C. polyneura
C1-1–C1-4TTZCTiantangzhai Scenic Spot, Lu’an, Anhui115.776731.129410404C. tschonoskii
Q1-1–Q1-3DPSQDapanshan National Nature Reserve, Pan’an, Zhejiang120.521628.971011313C. cordata
Table 2. Species information and GenBank IDs of ITS sequences obtained from the NCBI.
Table 2. Species information and GenBank IDs of ITS sequences obtained from the NCBI.
SpeciesGenBank ID
C. fangianaMG569958MG727560
Ostryopsis davidianaGQ250099KC412170
Corylus heterophyllaAF297351FJ011745
Betula platyphyllaAY761128FJ011778
Quercus acutissimaAF098428MN722083
Table 3. Nine ribotypes of C. tientaiensis identified based on 5 singleton variable sites and 3 parsimony information sites.
Table 3. Nine ribotypes of C. tientaiensis identified based on 5 singleton variable sites and 3 parsimony information sites.
RibotypesVariable Sites/bpNumber
62678605612614619641
R1T---T--A3
R2TA--T--A41
R3TACGT--A1
R4TA--G--A1
R5-A--TG-A1
R6TA--T---1
R7-A--T--A1
R8TA-GT--A2
R9TA--T-CA1
Table 4. Ribotype diversity, nucleotide diversity, and ribotype composition of C. tientaiensis.
Table 4. Ribotype diversity, nucleotide diversity, and ribotype composition of C. tientaiensis.
PopulationsRibotype Diversity (Hd)Nucleotide Diversity (π × 10−3)Ribotype NumberRibotype Composition and Number
SST0.331 ± 0.1140.7306R1 (1), R2 (23), R3 (1), R4 (1), R5 (1), R6 (1)
YTH0.000 ± 0.0000.0001R2 (2)
TTS0.507 ± 0.1400.8405R1 (2), R2 (12), R7 (1), R8 (1), R9 (1)
DPS0.400 ± 0.2370.5902R2 (4), R8 (1)
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Zhao, R.; He, Q.; Chu, X.; He, A.; Zhu, Z. Genetic Diversity of Carpinus tientaiensis Cheng, an Endemic and Critically Endangered Species in China, Based on ITS Sequences. Forests 2023, 14, 1600. https://doi.org/10.3390/f14081600

AMA Style

Zhao R, He Q, Chu X, He A, Zhu Z. Genetic Diversity of Carpinus tientaiensis Cheng, an Endemic and Critically Endangered Species in China, Based on ITS Sequences. Forests. 2023; 14(8):1600. https://doi.org/10.3390/f14081600

Chicago/Turabian Style

Zhao, Runan, Qianqian He, Xiaojie Chu, Anguo He, and Zunling Zhu. 2023. "Genetic Diversity of Carpinus tientaiensis Cheng, an Endemic and Critically Endangered Species in China, Based on ITS Sequences" Forests 14, no. 8: 1600. https://doi.org/10.3390/f14081600

APA Style

Zhao, R., He, Q., Chu, X., He, A., & Zhu, Z. (2023). Genetic Diversity of Carpinus tientaiensis Cheng, an Endemic and Critically Endangered Species in China, Based on ITS Sequences. Forests, 14(8), 1600. https://doi.org/10.3390/f14081600

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