Abstract
Paeonia japonica, distributed throughout Asia, is a traditional medicinal herb in Korea, with many potential beneficial effects including pain-relieving, anti-inflammatory, and anti-cancer activities. Despite its high pharmacological value, the genetic information on Paeonia japonica remains limited. In this study, the chloroplast genome of P. japonica was sequenced using next-generation sequencing (NGS) technology and genome and phylogeny were analyzed using multiple tools. The chloroplast genome of P. japonica was 152,731 bp in length with an inverted repeat region of 26,656 bp, including a large single-copy region of 84,389 bp and a small single copy region of 17,030 bp. The P. japonica chloroplast genome included 113 genes comprising 80 protein-coding genes, 27 tRNA, and 5 rRNA genes. Phylogenetic analysis indicated that P. japonica and P. obovata share a close evolutionary relationship.
Keywords: Chloroplast genome, Paeonia japonica, phylogenetic tree analysis
Paeonia japonica, belonging to the family Paeoniaceae, is distributed throughout Asia and its roots have long been used for medicinal purposes in South Korea. P. japonica is effective for treating pain (Zhang et al. 2014) and gynecological diseases (Xu et al. 2019). It is also known to have anti-inflammatory (Wu et al. 2010), vascular expansion (Park et al. 2009), and anti-tumor effects (Tan et al. 2020). Despite its pharmacological importance, limited genetic information about P. japonica is available. Thus, in this study, we identified the sequence of the complete chloroplast genome sequence of P. japonica using next-generation sequencing techniques and then performed phylogenetic analysis.
Samples of P. japonica were collected from Goesan-gun (Chungcheongbuk-do, South Korea, 36°45'47.7"N 127°53'24.8"E) and Yangju-si (Gyeonggi-do, South Korea, 37°48'15.1"N 127°01'32.6"E). Voucher specimens (TKMII-23-2) were deposited to the Medicinal Crops Seed Supply Center of the National Institute for Korean Medicine Development. Chloroplast DNA was extracted using the DNeasy Plant Mini Kit (Qiagen, Germany) and DNA library was created using EBNext Ultra II DNA Library Prep Kit for Illumina (NEB). The generated library used TapeStation HSD5000 (Agilent) to analyze the condition and determine the suitability of NGS analysis. The prepared library sequenced using the Illumina HiSeq 2500 Platform with 151 bp paired-end reads at Genotech (Daejeon, Korea). About 3.25 Gb raw reads (9639.196 bp) were generated and assembled using NOVOPlasty v2.6.7 (Dierckxsens et al. 2017). Gene annotation was performed using GeSeq (Tillich et al. 2017) and CPGAVAS2 (Shi et al. 2019). When performing assembly and annotation, the Paeonia lactiflora chloroplast genome (GenBank accession: NC_040983.1) was used as a reference genome. The annotated chloroplast genome sequence was submitted to the NCBI GenBank database under the accession number MT821944. The circular genome annotation map was drawn using OGDRAW v1.3.1 (Greiner et al. 2019). Next, to determine the phylogenetic position of P. japonica, 19 complete chloroplast genomes belonging to the Paenoniaceae and 4 outgroup sequences were aligned using MAFFT v7 (Katoh and Standley 2013). A maximum likelihood phylogenetic tree was generated using MEGA X (Kumar et al. 2018) with 1000 bootstrap replicates.
The complete chloroplast genome of P. japonica was 152,731 bp in length and included a pair of inverted repeats (IRa and IRb) spanning 25,656 bp, separated by a large single copy region (LSC) of 84,389 bp and a small single copy region (SSC) of 17,030 bp. Overall, the GC content of the P. japonica chloroplast genome was 38.4%, and that of the LSC and SSC regions was 36.7% and 32.7%, respectively. In total, the P. japonica chloroplast genome included 113 genes. These included 80 protein-coding genes, 27 tRNA genes, and 5 rRNA genes. The maximum likelihood phylogenetic tree showed that Paeonia species were clustered together (Figure 1), and that P. japonica was closely related to P. obovata. This decoded chloroplast genome sequence of P. japonica will be useful for studying species conservation strategies and pharmacological efficacy in the future.
Figure 1.
The ML phylogenetic tree constructed with total 23 chloroplast genome sequences. (Bootstrap replicates = 1000). All the sequences were downloaded from GenBank.
Acknowledgements
We thank Dr. S. H. Cha and S.W. Lee for assisting with the nucleotide data analysis of plant materials.
Funding Statement
This work was supported by the Korean Medicinal Herb-Based Business of the Korean Traditional Resource [Ministry of Health and Welfare, Republic of Korea].
Disclosure statement
The authors declare that there are no conflicts of interest.
Data availability statement
The data generated in this study are publicly available at NCBI GenBank (https://www.ncbi.nlm.nih.gov/genbank/), under reference number MT821944.
References
- Dierckxsens N, Mardulyn P, Smits G.. 2017. NOVOPlasty: de novo assembly of organelle genomes from whole genome data. Nucleic Acids Res. 45(4):e18. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Greiner S, Lehwark P, Bock R.. 2019. OrganellarGenomeDRAW (OGDRAW) version 1.3.1: expanded toolkit for the graphical visualization of organellar genomes. Nucleic Acids Res. 47(W1):W59–W64. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Katoh K, Standley DM.. 2013. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 30(4):772–780. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kumar S, Stecher G, Li M, Knyaz C, Tamura K.. 2018. MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol. 35(6):1547–1549. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Park SG, Lee MJ, Jung HJ, Lee HS, Kim H, Na ST, Park SD, Park WH.. 2009. Preventive effects of peony root extracts on oxidative stress, thrombosis and atherosclerosis. J Kor Oriental Med. 30(2):88–103. [Google Scholar]
- Shi L, Chen H, Jiang M, Wang L, Wu X, Huang L, Liu C.. 2019. CPGAVAS2, an integrated plastome sequence annotator and analyzer. Nucleic Acids Res. 47(W1):W65–W73. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tan YQ, Chen HW, Li J, Wu QJ.. 2020. Efficacy, chemical constituents, and pharmacological actions of Radix Paeoniae Rubra and Radix Paeoniae Alba. Front Pharmacol. 11:1054. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tillich M, Lehwark P, Pellizzer T, Ulbricht-Jones ES, Fischer A, Bock R, Greiner S.. 2017. GeSeq – versatile and accurate annotation of organelle genomes. Nucleic Acids Res. 45(W1):W6–W11. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wu SH, Wu DG, Chen YW.. 2010. Chemical constituents and bioactivities of plants from the genus Paeonia. Chem Biodivers. 7(1):90–104. [DOI] [PubMed] [Google Scholar]
- Xu Y, Li X, Chen T, Qu YK, Zheng HX, Zhang ZJ, Zhao Y, Lin N.. 2019. Radix Paeoniae Alba increases serum estrogen level and up-regulates estrogen receptor expression in uterus and vagina of immature/ovariectomized mice . Phytother Res. 33(1):117–129. [DOI] [PubMed] [Google Scholar]
- Zhang Y, Zhou R, Zhou F, Cheng H, Xia B.. 2014. Total glucosides of peony attenuates 2,4,6-trinitrobenzene sulfonic acid/ethanol-induced colitis in rats through adjustment of TH1/TH2 cytokines polarization. Cell Biochem Biophys. 68(1):83–95. [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The data generated in this study are publicly available at NCBI GenBank (https://www.ncbi.nlm.nih.gov/genbank/), under reference number MT821944.