The complete chloroplast genome of Agrimonia pilosa Ledeb. isolated in Korea (Rosaceae): investigation of intraspecific variations on its chloroplast genomes.

The complete chloroplast genome of Agrimonia pilosa Ledeb. isolated in Korea is 155,125 bp long (GC ratio is 36.9%) and has four subregions: 84,458 bp of large single copy (34.9%) and 18,737 bp of small single copy (30.4%) regions are separated by 25,965 bp of inverted repeat (42.6%) regions including 129 genes (84 protein-coding genes, eight rRNAs, and 37 tRNAs). 258 SNPs and 542 INDELs were identified as intraspecific variations against the partial genome (KY419942). Phylogenetic trees show that our chloroplast genome was clustered with the previous A. pilosa chloroplast genome.

Agrimonia pilosa Ledeb., which is a perennial herb in tribe Sanguisorbeae of Rosaceae, is primarily distributed over the Korean peninsula, Japan, China, Siberia, and Eastern Europe (Chung and Kim 2000; Seo et al. 2017). There are neighbor species in Korea, such as A. coreana and A. nipponica, considered as a variety of A. pilosa for a while (Chung and Kim 2000). Its root and aerial parts have been used as Traditional Chinese Medicines for hemostatic, antimalarial, and anti-dysenteric treatments (Park et al. 2004). Its partial chloroplast genome was uncovered from the individual which might be isolated in China (KY419942; Zhang et al. 2017) and its complete chloroplast genome was also published (Liu et al. 2020) but the sequence is not available as of May 2020.

Total DNA of A. pilosa isolated from Mt. Kwangdeok, Gangwon-do, Republic of Korea, was extracted from fresh leaves using a DNeasy Plant Mini Kit (QIAGEN, Hilden, Germany). The voucher was deposited in InfoBoss Cyber Herbarium (IN; Heo K-I, IB-01030; 38°06′21.7″N, 127°26′37.6″E). Genome was sequenced using HiSeqX at Macrogen Inc., Korea, and de novo assembly and confirmation were performed using Velvet 1.2.10 (Zerbino and Birney 2008), SOAPGapCloser 1.12 (Zhao et al. 2011), BWA 0.7.17 (Li 2013), and SAMtools 1.9 (Li et al. 2009). Geneious R11 11.0.5 (Biomatters Ltd., Auckland, New Zealand) was used for annotation based on A. pilosa chloroplast (KY419942).

Chloroplast genome of A. pilosa (GenBank accession is MT415946) is 155,125 bp (GC ratio is 36.9%) and has four subregions: 84,458 bp of large single copy (LSC; 34.9%) and 18,737 bp of small single copy (SSC; 30.4%) regions are separated by 25,695 bp of inverted repeat (IR; 42.6%). It contains 129 genes (84 protein-coding genes, eight rRNAs, and 37 tRNAs); 17 genes (seven protein-coding gene, four rRNAs, and six tRNAs) are duplicated in IR regions.

Due to lack of available complete chloroplast genome, partial chloroplast was used for identifying intraspecific variations: 258 SNPs and 542 INDELs were identified from aligned LSC, SSC, and IRb regions. Number of intraspecific variations is relatively large: several plant species, such as Goodyera schlechtendaliana (Oh et al. 2019a, 2019b), Gastrodia elata (Kang et al. 2020; Park, Suh, et al. 2020), Pyrus ussuriensis (Cho et al. 2019), Camellia japonica (Park, Kim, et al. 2019), Selaginella tamariscina (Park, Kim, et al. 2020), Marchantia polymorpha subsp. ruderalis (Kwon et al. 2019), displayed larger number of variations.

Thirty-eight chloroplast genomes including two A. pilosa chloroplasts were used for constructing bootstrapped maximum-likelihood neighbor-joining trees using MEGA X (Kumar et al. 2018) after aligning whole chloroplast genomes using MAFFT 7.450 (Katoh and Standley 2013) with adjusting three chloroplast genomes. Phylogenetic trees show that our chloroplast genome was clustered with the previous partial genome (Zhang et al. 2017; Figure 1). In addition, the topology of the trees agrees with the previous study (Zhang et al. 2017; Figure 1). Our chloroplast genome can be utilized to understand phylogenetic relationship of neighbor Agrimonia species in Korea, A. coreana and A. nipponica, after their complete chloroplast genomes are available.

Figure 1.

Neighbor joining (bootstrap repeat is 10,000) and maximum-likelihood (bootstrap repeat is 1,000) phylogenetic trees of ten Rosa chloroplast genomes and three outgroup species: Agrimonia pilosa (MT415946 in this study and KY419942; partial genome; Zhang et al. 2017), Acaena pinnatifida (KY419984; partial genome; Zhang et al. 2017), Bencomia exstipulata (NC_039924), Bencomia moquiniana (KY420023; partial genome; Zhang et al. 2017), Bencomia sphaerocarpa (KY419986; partial genome; Zhang et al. 2017), Cliffortia repens (KY419983; partial genome; Zhang et al. 2017), Dendriopoterium menendezii (KY419966; partial genome; Zhang et al. 2017), Fragaria x ananassa cultivar Benihoppe (NC_035961; Cheng et al. 2017), Hagenia abyssinica (KX008604; Gichira et al. 2017) and KY420026; partial genome; Zhang et al. 2017), Leucosidea sericea (KY419929; partial genome (Zhang et al. 2017)), Margyricarpus pinnatus (KY419972; partial genome; Zhang et al. 2017), Polylepis australis (KY419989; partial genome; Zhang et al. 2017), Polylepis reticulata (KY419921; partial genome; Zhang et al. 2017), Polylepis sp. SCZ-2017 (KY419992; partial genome; Zhang et al. 2017), Potentilla centigrana (NC_041209; Park et al. 2019a), Potentilla freyniana (NC_041210; Park et al. 2019b), Duchesnea chrysantha (NC_041199; Park, Heo, et al. 2019), Duchesnea indica (NC_041178; Heo, Kim, et al. 2019), Potentilla stolonifera var. quelpaertensis (NC_044418; Heo, Park, et al. 2019), Poterium spinosum (KY419948; partial genome; Zhang et al. 2017), Rosa chinensis var. spontanea (NC_038102; Jian et al. 2018), Rosa multiflora (NC_039989; Jeon and Kim 2019), Rosa roxburghii (NC_032038; Wang et al. 2018), Rosa rugosa (NC_044094; Kim et al. 2019), Rosa angusta (NC_044126; Kim et al. 2019), Rubus crataegifolius (NC_039704; Yang et al. 2017), Sanguisorba filiformis (NC_044693 (Meng et al. 2018) and KY419920; partial genome; Zhang et al. 2017), Sanguisorba officinalis (NC_044694 (Meng et al. 2018) and MK696193), Sanguisorba sitchensis (NC_044691; Meng et al. 2018), Sanguisorba tenuifolia var. alba (NC_044692; Meng et al. 2018), Sanguisorba tenuifolia (NC_042223 (Park et al. 2018) and MK696195), Spenceria ramalana (KY419995; partial genome; Zhang et al. 2017), Sanguisorba stipulate (MK696195; partial genome; Zhang et al. 2017). Phylogenetic tree was drawn based on maximum-likelihood tree. The numbers above branches indicate bootstrap support values of maximum-likelihood and neighbor-joining phylogenetic tree, respectively.