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Article

DNA Barcodes for Wood Identification of Anatomically Similar Species of Genus Chamaecyparis

by
Minjun Kim
1,2,†,
Seokhyun Im
1,2,† and
Tae-Jong Kim
1,2,*
1
Department of Forest Products and Biotechnology, Kookmin University, Seoul 02707, Republic of Korea
2
Forest Carbon Graduate School, Kookmin University, Seoul 02707, Republic of Korea
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Forests 2024, 15(7), 1106; https://doi.org/10.3390/f15071106
Submission received: 22 May 2024 / Revised: 20 June 2024 / Accepted: 23 June 2024 / Published: 27 June 2024
(This article belongs to the Section Wood Science and Forest Products)
Browse Figures
Versions Notes

Abstract

:
The genus Chamaecyparis comprises seven species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis (Callitropsis nootkatensis), C. obtusa, C. pisifera, and C. thyoides). Accurate species identification is necessary for proper use and economic value of wood. Species identification of woods is generally based on anatomical analysis; however, C. obtusa and C. pisifera wood have similar microscopic morphology, which makes species identification impossible. Therefore, the molecular identification of species in wood of the genus Chamaecyparis is required. In this study, six candidate DNA barcode genes (trnP-GGG, ycf1b, clpP, accD, ycf2, and rps16) in the chloroplast of Chamaecyparis were identified with nucleotide diversity values higher than the arbitrary value of 0.02. Each gene was evaluated for species identification using phylogenetic analysis by genes registered at NCBI (42 sequences each for trnP-GGG, ycf1b, clpP, accD, and ycf2, and 50 sequences for rps16). The genes trnP-GGG, clpP, and rps16 could not be distinguished between C. pisifera and C. formosensis. However, ycf1b, accD, and ycf2 could be distinguished between all Chamaecyparis species. These results suggest the use of the chloroplast genes ycf1b, accD, and ycf2 as DNA barcodes for species identification in Chamaecyparis, including C. obtusa and C. pisifera, based on the reported genetic information to date.

1. Introduction

A survey in 2012 found that 15–30% of forests were being illegally logged worldwide [1]. Unplanned logging causes landslides and flooding, which adversely affect the safety and economy of local communities [2]. Illegal logging also adversely affects the environment, as forests serve as carbon reservoirs as well as natural habitats for many animals and plants [2,3]. According to the 2006 World Bank data, illegal logging causes an estimated annual loss of USD 1.5 billion, with losses in the legal forest industry accounting for more than 60% of the total [3,4]. Many countries, including the United States, Australia, and countries in Europe, have banned the import and trade of illegal timber [5,6]. Accurate wood identification is critical for the successful enforcement of regulations against the illegal timber trade [6]. Wood identification can also help protect forests by controlling the trade of wood obtained from endangered species or forests in need of protection [6]. DNA barcoding for timber species identification can authenticate their origin [7]. The Forest Stewardship Council (FSC), which tracks and manages the harvesting, processing, and distribution of timber, argues that tracing the origin of timber helps to prevent illegal logging and can also help to ensure a sustainable forest [8,9]. Consumers can access wood based on species-specific characteristics by using wood identification [10].
The genus Chamaecyparis is mainly found in East Asia and North America and their wood is used as high-end building and furniture materials [11,12,13]. Chamaecyparis comprises seven species, including C. formosensis, C. hodginsii, C. lawsoniana, Callitropsis nootkatensis (homotypic synonym: C. nootkatensis), C. obtusa, C. pisifera, and C. thyoides [14,15]. The mixed forests of subtropical eastern Asia are inhabited by the genus Chamaecyparis, especially C. lawsoniana [11,16]. C. formosensis is used for furniture due to its high wood quality, aroma, and durability [17]. C. lawsoniana is widely planted for landscaping in North America and Europe, where it has ecological and economical value [18]. Extracts of C. obtusa are used in medicinal preparations for their antifungal and anti-inflammatory properties [19,20]. C. formosensis and C. obtusa are threatened by illegal logging in Taiwan [21,22]. The conservation statuses of C. formosensis, C. hodginsii, C. lawsoniana, and C. obtusa are endangered, vulnerable, near threatened, and near threatened, respectively (https://threatenedconifers.rbge.org.uk/taxonomy/cupressaceae/p2 (accessed on 25 February 2024)). C. lawsoniana is found in a limited region and requires protection from logging and root rot disease [11]. Therefore, each species of Chamaecyparis has different reasons for accurate species identification, depending on the situation.
Wood species are primarily identified by observing anatomical features under a microscope [4,5,23,24,25,26]. However, C. obtusa and C. pisifera have similar pit numbers, pit size, pit type, ray frequency, and ray height, making it difficult to distinguish them based on anatomical characteristics alone [27]. DNA barcoding technology is an alternative method that can be used to distinguish anatomically similar species [28,29]. Using genes from chloroplasts to identify wood species has many advantages. The presence of a large number of identical chloroplasts in a single cell makes it easier to obtain analyzable chloroplast genes from old, processed wood. In addition, chloroplast DNA multiplies by binary fission and is inherited only from the maternal line, so there is no mixing of genes through fertilization, providing genes that can be used for species identification. Previous studies showed that phylogenetic analysis of the genus Chamaecyparis using matK, which is a chloroplast DNA barcode commonly used to distinguish plants, failed to distinguish between C. formosensis and C. pisifera [30]. DNA barcoding using the internal transcribed spacer region, which is generally used to classify eukaryotic cells, also failed to distinguish between C. lawsoniana and C. obtusa [31,32]. Further, petG-trnP and trnV cannot be used to distinguish between C. formosensis, C. lawsoniana, and C. obtusa [14].
In the present study, we propose new DNA barcodes that can accurately identify all seven species of Chamaecyparis, including species that are anatomically indistinguishable. For this purpose, all complete chloroplast genome sequences of the genus Chamaecyparis were obtained from the National Center for Biotechnology Information (NCBI), and genes with high variation were selected. A phylogenetic analysis of the selected genes was performed to evaluate their functionality as DNA barcodes. Accurate identification of all species in the genus Chamaecyparis provides an opportunity for the many benefits mentioned above.

2. Materials and Methods

2.1. Measurement of Nucleotide Diversity

The nine chloroplast genomes (NCBI accession numbers: LC522362 (C. formosensis), NC_034943 (C. formosensis), KX832623 (C. hodginsii), MG269834 (C. hodginsii), KX832622 (C. lawsoniana), LC529363 (C. obtuse), MT258872 (C. obtusa), MT334621 (C. pisifera), and NC_057503 (C. pisifera)), for which complete sequences have been reported in the genus Chamaecyparis, were collected from the NCBI database and aligned using the ClustalW program [33]. Nucleotide diversity was calculated using the DnaSP (ver. 6.0) software with 100 bp of window length and 50 bp of step size [34].

2.2. Gene Alignment and Phylogenetic Analysis

Gene alignments and phylogenetic analysis were performed as described in our previous study [35]. Briefly, gene sequences of accD, clpP, trnP-GGG, ycf1b, ycf2, and rps16, with a query coverage of more than 90% for the genus Chamaecyparis, were collected and aligned using the ClustalW program [33]. Forty-two genes for each accD, clpP, trnP-GGG, ycf1b, and ycf2, and fifty genes for rps16 were obtained from the NCBI database. The NCBI accession numbers of the genes used are listed in each Figures, showing the results of the phylogenetic analysis, and in the Supplementary Material Figures. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method [36]. trnP-GGG from Juniperus chinensis; ycf1b, clpP, accD, and ycf2 from Thuja occidentalis; and rps16 from Calocedrus formosana were used as an outgroup. Alignment results were displayed using BioEdit (ver. 7.7.1) to present the sequence alignment [37].

3. Results and Discussion

3.1. Evaluation of Nucleotide Diversity of Chloroplast Genomes within Chamaecyparis

The nucleotide diversity of each gene was assessed using the DnaSP (ver. 6.0) software with the nine reported complete chloroplast genomes of the genus Chamaecyparis (NCBI accession numbers: LC522362 (C. formosensis), NC_034943 (C. formosensis), KX832623 (C. hodginsii), MG269834 (C. hodginsii), KX832622 (C. lawsoniana), LC529363 (C. obtuse), MT258872 (C. obtusa), MT334621 (C. pisifera), and NC_057503 (C. pisifera)) (Figure 1). Intergenic spacer sequences can exhibit high variation due to inversions and insertions within sequences of the same species, making species identification difficult [38]. Therefore, the nucleotide diversity of all genes in the chloroplasts was evaluated. Nucleotide diversity is an indicator of the genetic variation in a gene across species, and genes with high diversity can be used as DNA barcodes [39]. Six genes, trnP-GGG, ycf1, clpP, accD, ycf2, and rps16, with arbitrary nucleotide diversity (pi) values > 0.02 were selected for functional evaluation as DNA barcodes.
trnP-GGG, a transfer RNA coding for GGG to glycine [40], had the highest pi value. A previous study similarly reported that trnP-GGG in the Cupressaceae family has a high pi value [41]. ycf1, which had the second-highest pi value, often shows a high pi value among species and has been used in previous studies to identify members of the genus Pinus [42] and the family Orchidaceae [43]. In the genus Chamaecyparis, ycf1 is a long gene with a nucleotide sequence of approximately 7000 bp, which exceeds the optimal length for use as a DNA barcode. Therefore, we used one of the small regions of ycf1, ycf1b, which has previously shown high efficiency as a DNA barcode [44]. clpP encodes a subunit of ATP-dependent protease [45]. clpP has been proposed as a DNA barcode for 27 species of the family Actinidiaceae; however, it cannot accurately distinguish all species [46]. accD encodes the acetyl-CoA carboxylase subunit D [47]. accD is used in combination with other genes to classify the genus Hexachlamys; however, two of the four species cannot be accurately identified [48]. ycf2, which encodes an FstH-like protein [49], shows the second-highest gene diversity among eight genes (matK, rbcL, rpl20-rps18, trnH-psbA, trnL-trnF, trnV, ycf1, and ycf2) within the genus Pinus, thereby demonstrating its potential as a DNA barcode [50]. rps16 encodes a ribosomal protein [51], and its functionality as a DNA barcode for identifying species of the genus Bupleurum was evaluated in conjunction with an internally transcribed spacer; however, it cannot accurately identify species [52].

3.2. Phylogenetic Analysis of Five Species of Chamaecyparis Using trnP-GGG

trnP-GGG sequences of only five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) are available in the NCBI database. All 42 reported trnP-GGG sequences from the genus Chamaecyparis, including genes from the complete chloroplast genome presented in Figure 1, were used to evaluate their functionality as a DNA barcode for species identification (Figure 2). Phylogenetic analysis could not accurately distinguish between C. formosensis and C. pisifera. When the gene sequences were aligned (Supplementary Material Figure S1), there were seven mismatches out of 72 bases. trnP-GGG showed the highest nucleotide diversity among chloroplast genes belonging to Chamaecyparis. However, the trnP-GGG genes of two indistinguishable species (C. formosensis and C. pisifera) had identical nucleotide sequences. A previous study suggested that many mutations in trnP-GGG of Tortula ruralis and Physcomitrella patens suggest the possibility of a pseudogene [53].

3.3. Phylogenetic Analysis of Five Species of Chamaecyparis Using ycf1b

We investigated whether ycf1b can classify five species of Chamaecyparis (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) based on phylogenetic analysis (Figure 3). In addition to the nine reported complete chloroplast genes presented in Figure 1, an additional thirty-three ycf1b sequences were used. Since all five species form an independent lineage group, ycf1b can be used as a DNA barcode for species identification in the genus Chamaecyparis. The length of the aligned ycf1b nucleotide sequence was 1469 bases, and the number of non-identical nucleotide sequences was 130 bases (Supplementary Material Figure S2). ycf1b could be used to distinguish between C. formosensis and C. pisifera. ycf1b can be used to identify 71.87% of 391 land plant species [54]. In the present study, ycf1b could also be used to accurately identify species of Chamaecyparis. Since the sequence of ycf1b for C. nootkatensis was not available, it was not included in this evaluation. ycf1b could be used to distinguish C. obtusa from other species; however, it could not be used to distinguish the subspecies of C. obtusa.

3.4. Phylogenetic Analysis of Six Species of Chamaecyparis Using clpP

Since clpP gene sequences of C. nootkatensis have been published, we evaluated the functionality of clpP as a DNA barcode in six species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) of Chamaecyparis (Figure 4). In addition to the nine reported complete chloroplast genes presented in Figure 1, 33 clpP sequences were analyzed. The length of the clpP for alignment was 540 bases, and the number of non-identical nucleotides was 45 bases (Supplementary Material Figure S3). The clpP nucleotide sequences of C. formosensis and C. pisifera were identical. In contrast, there was a mismatch of a single base among the 540-base alignment between C. lawsoniana and C. obtusa, making accurate species identification impossible. Previously, mutations in the 14th–33rd bases region of clpP have been shown to be important for the identification of 27 Actinidiaceae species [46]. In Chamaecyparis species, clpP had a mutation starting from the 103rd position, excluding C. nootkatensis. clpP has been proposed as a DNA barcode for 27 species of Actinidiaceae in a previous study; however, it cannot accurately identify all species [46].

3.5. Phylogenetic Analysis of Six Species of Chamaecyparis Using accD

We investigated whether accD could be used as a DNA barcode for six species of Chamaecyparis (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) (Figure 5). In addition to the nine reported complete chloroplast genes presented in Figure 1, 33 accD sequences were analyzed. All six species formed an independent lineage. The nucleotide sequence of accD for alignment was 2208 bases, and the number of non-identical nucleotides was 299 bases (Supplementary Material Figure S4). This result was inconsistent with results presented in Figure 1, where accD showed the fourth-highest nucleotide diversity; this was attributed to additional accD sequences for C. nootkatensis. The number of non-identical nucleotides, excluding those of C. nootkatensis, was 162 bases. accD could be used to distinguish between two subspecies, C. obtusa var. obtusa and C. obtusa var formosana. The accD sequence of C. obtusa had six mutations when compared to accD sequences of other species. Among them, three nucleotides (34th, 1098th, and 1544th) were identical across subspecies (Supplementary Material Figure S4). The accD gene had sufficient nucleotide diversity to identify six species. Amino acid repeats have been found in accD, and the number of repetitions differs depending on the species [55]. In Chamaecyparis, a region with repeats of 10 amino acid sequences was found at the beginning.

3.6. Phylogenetic Analysis of Six Species of Chamaecyparis Using ycf2

ycf2 was evaluated as a DNA barcode for six species of Chamaecyparis (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) (Figure 6). In total, 42 reported ycf2 sequences were analyzed. All six species could be identified, and nucleotide diversity was present within C. formosensis, forming six subspecies lineage groups. Unlike other genes, ycf2 had variations in six nucleotides (3616th, 3629th, 3654th, 3655th, 3689th, and 3690th) within C. formosensis (Supplementary Material Figure S5). The nucleotide sequence of ycf2 for alignment was 6748 bases, and the non-identical nucleotide was 720 bases (Supplementary Material Figure S5). Like accD, ycf2 from C. nootkatensis, for which a complete genome sequence is unavailable, increased the nucleotide diversity compared to that presented in Figure 1. The non-identical nucleotide sequence, excluding C. nootkatensis sequences, was 396 bases. ycf2 could be used to distinguish C. obtusa from other species as well as its subspecies.

3.7. Phylogenetic Analysis of Five Species of Chamaecyparis Using rps16

We investigated whether rps16 could be used as a DNA barcode for five species of Chamaecyparis (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) (Figure 7). In addition to the nine reported complete chloroplast genes presented in Figure 1, 41 rps16 sequences were analyzed. The total nucleotide sequence of rps16 was 374 bases, and the non-identical nucleotide was 24 bases (Supplementary Material Figure S6). C. pisifera and C. formosensis are phylogenetically similar, with a chloroplast DNA divergence of 0.57% [11]. rps16 also had only one mutation at the 170th position in the alignment between C. formosensis and C. pisifera. Therefore, it could not be used to accurately identify the species.

4. Conclusions

In this study, we proposed three genes, ycf1b, accD, and ycf2, as DNA barcodes for the molecular and phylogenetic classification of the genus Chamaecyparis, which is difficult to distinguish by species based on anatomical characteristics alone. To select DNA barcodes for species identification, six candidate genes with high nucleotide diversity in chloroplast genes were selected, and the accuracy of species identification was evaluated via phylogenetic analysis. Based on all available ycf1b, accD, and ycf2 sequences in the NCBI nucleotide database on 1 March 2024, these genes allowed the identification of Chamaecyparis species. In contrast, trnP-GGG, clpP, and rps16 failed to accurately distinguish C. pisifera and C. formosensis. Therefore, using ycf1b, accD, and ycf2 in phylogenetic analysis can accurately identify species of the genus Chamaecyparis. This study was analyzed based on genes reported to date, and new useful DNA barcodes may be proposed by the accumulation of additional genetic data; however, it can be used as an auxiliary method to clearly identify species in the genus Chamaecyparis when anatomical species identification is not possible. The method of exploring DNA barcodes in this study can be utilized as a method that can be used for taxonomic identification of subspecies.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/f15071106/s1, Figure S1: Sequence alignment of 42 trnP-GGG sequences in the genus Chamaecyparis. The same base with the NC034943 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page; Figure S2: Sequence alignment of 42 ycf1b sequences in the genus Chamaecyparis. The same base with the KP089387 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page.; Figure S3: Sequence alignment of 42 clpP sequences in the genus Chamaecyparis. The same base with the KX832622 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page.; Figure S4: Sequence alignment of 42 accD sequences in the genus Chamaecyparis. The same base with the NC034943 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page.; Figure S5: Sequence alignment of 42 ycf2 sequences in the genus Chamaecyparis. The same base with the LC522086 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page.; Figure S6: Sequence alignment of 50 rps16 sequences in the genus Chamaecyparis. The same base with the OK616152 represent as “.” and the only different bases are presented. The species names of the sequences are presented in the last page.

Author Contributions

Conceptualization, M.K. and T.-J.K.; methodology, M.K. and S.I.; software, S.I.; validation, M.K., S.I. and T.-J.K.; formal analysis, S.I. and T.-J.K.; investigation, S.I.; resources, T.-J.K.; data curation, S.I. and T.-J.K.; writing—original draft preparation, S.I.; writing—review and editing, T.-J.K.; visualization, S.I.; supervision, T.-J.K.; project administration, T.-J.K.; funding acquisition, M.K. and S.I. All authors have read and agreed to the published version of the manuscript.

Funding

This study was funded by the R&D Program for Forest Science Technology (Project No. RS-2024-00358413) of the Korea Forestry Promotion Institute (Korea Forest Service, Forest Policy Division, Seoul, Republic of Korea).

Data Availability Statement

The datasets used/analyzed during the current study are available from the corresponding author upon reasonable request.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. Analysis of nucleotide diversity values of genes in nine complete chloroplast genomes of the genus Chamaecyparis (C. formosensis (LC522362, NC_034943), C. hodginsii (KX832623, MG269834), C. lawsoniana (KX832622), C. obtusa (LC529363, MT258872), and C. pisifera (MT334621, NC_057503)) using DnaSP (ver. 6.0) software. Pi, nucleotide diversity value. Due to the large number of genes, it is presented in two graphs.
Figure 1. Analysis of nucleotide diversity values of genes in nine complete chloroplast genomes of the genus Chamaecyparis (C. formosensis (LC522362, NC_034943), C. hodginsii (KX832623, MG269834), C. lawsoniana (KX832622), C. obtusa (LC529363, MT258872), and C. pisifera (MT334621, NC_057503)) using DnaSP (ver. 6.0) software. Pi, nucleotide diversity value. Due to the large number of genes, it is presented in two graphs.
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Figure 2. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using trnP-GGG. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 2. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using trnP-GGG. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Figure 3. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using ycf1b. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 3. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using ycf1b. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Figure 4. Phylogenetic analysis of six Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using clpP. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 4. Phylogenetic analysis of six Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using clpP. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Figure 5. Phylogenetic analysis of six Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using accD. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 5. Phylogenetic analysis of six Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using accD. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Figure 6. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using ycf2. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 6. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. nootkatensis, C. obtusa, and C. pisifera) using ycf2. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Figure 7. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using rps16. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
Figure 7. Phylogenetic analysis of five Chamaecyparis species (C. formosensis, C. hodginsii, C. lawsoniana, C. obtusa, and C. pisifera) using rps16. Phylogenetic analysis was performed with 1000 bootstraps using the Kimura two-parameter model in the MEGA (ver. 11.0) software using the maximum likelihood method.
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Kim, M.; Im, S.; Kim, T.-J. DNA Barcodes for Wood Identification of Anatomically Similar Species of Genus Chamaecyparis. Forests 2024, 15, 1106. https://doi.org/10.3390/f15071106

AMA Style

Kim M, Im S, Kim T-J. DNA Barcodes for Wood Identification of Anatomically Similar Species of Genus Chamaecyparis. Forests. 2024; 15(7):1106. https://doi.org/10.3390/f15071106

Chicago/Turabian Style

Kim, Minjun, Seokhyun Im, and Tae-Jong Kim. 2024. "DNA Barcodes for Wood Identification of Anatomically Similar Species of Genus Chamaecyparis" Forests 15, no. 7: 1106. https://doi.org/10.3390/f15071106

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