The complete chloroplast genome of Pleione maculata, an orchid with important ornamental value and medicinal value

Abstract Pleione maculata is an epiphytic orchid with significant ornamental value and medicinal value. Here, we report the first complete chloroplast genome of P. maculata. The circular genome was 158,394 bp in length and consisted of a pair of inverted repeats (IR 26,646 bp), which were separated by a large single copy region (LSC 86,603 bp) and a small single copy region (SSC 18,499 bp). It contained 135 genes, including 89 protein-coding genes, 38 tRNAs and 8 rRNAs. Phylogenetic analysis of cp genomes from 41 species of Orchidaceae revealed that all species of Pleione formed one monophyletic clade and P. maculata was located at the base of the genus with high bootstrap values (≥99.1%).

Pleione maculata; Illumina sequencing; complete chloroplast genome; phylogenetic analysis The genus Pleione D. Don (Orchidaceae), comprising about 33 speices (7 natural hybrid species) of terrestrial, lithophytic or epiphytic orchids, is widely distributed in China, Vietnam, India, Bhutan, Nepal, Thailand, and Myanmar (Chen et al. 2009;Govaerts et al. 2016). Pleione are fairly popular ornamental plants in Europe and the USA and widely used as traditional medicine in Asian countries (Teoh 2016). Pleione bulbocodioides (Franchet) Rolfe and P. yunnanensis (Rolfe) Rolfe are also the original plant of traditional Chinese medicine Sh an C ı G u. The pseudobulb was called commonly 'ice ball', which has the functions of clearing heat, detoxifying, and resolving phlegm (You et al. 2011). Pleione is also a world-famous ornamental, with showy, color-rich and leafless flowers in flowering period (Zhang et al. 2020).
Pleione maculata (Lindley) Lindley & Paxton, distributed in Bhutan, India, Myanmar, Nepal, Thailand and China between 600 and 1600 meters above sea level, is an epiphytic herb growing on tree trunks and mossy rocks in broad-leaved forests (Chen et al. 2009). Pseudobulbs of P. maculata, has commonly been applied in the northeast India for the treatment of cuts, wound, or liver complaints (Simpli et al. 2018). In the wild, P. maculata is one of the most important parents for a succession of hybrids (Gravendeel et al. 2004;Chen et al. 2009). Although Pleione have high ornamental value and medicinal value, there are a few reports on the chloroplast genome. So far, all species with cp genomes reported belong to the Sect. Humiles, such as, P. bulbocodioides  (Chen et al. 2019). In this study, we assembled and characterized the complete chloroplast of P. maculata to provide a better understanding of the phylogeny and genetics of genus Pleione.
Fresh leaves of P. maculata were collected from the individual growing in the greenhouse of Yunnan Normal University, Kunming, China (24 52'1.41'N, 102 51'19.39'E), and voucher specimen (OP-001) deposited at Herbarium of Yunnan Normal University (YNUB). Total genomic DNA was extracted from fresh leaves using a modified CTAB method (Allen et al. 2006) and sequenced by the Illumina Hiseq 2000 sequencing platform (Illumina, CA, USA) at Novogene (Beijing, China). In total, 3.44 GB of raw data with 11,464,366 raw reads was obtained. Raw reads were filtered by NGS QC Toolkit (Patel et al. 2012). The plastome was de novo assembled using NOVOPlasty (Dierckxsens et al. 2017). After assembled, the genome was automatically annotated using DOGMA (Wyman et al. 2004), then adjusted with Geneious Prime 2020.0.3 (https://www.geneious.com) and submitted to GenBank with accession number MW699846.
To clarify the phylogenetic position of P. maculata, 41 published chloroplast genomes from Orchidaceae were aligned by using MAFFT 7.308 (Katoh and Standley 2013) with Goodyera fumata (GenBank accession KJ501999) and Ludisia discolor (GenBank accession NC030540) as outgroups. Maximum-likelihood (ML) tree was constructed using RAxML 8.2.11 (Stamatakis 2014) with the GTR þ G nucleotide substitution model. The branch supports were computed with 1000 bootstrap replicates. The phylogenetic tree showed that six species of Pleione formed one monophyletic clade with 100% bootstrap value ( Figure 1) and P. maculata was located at the base of the genus with high bootstrap value (99.1%). The complete chloroplast genome of P. maculata will provide useful resource for identification, conservation, and utilization of this valuable species. Moreover, it will be helpful to better understand the phylogeny of genus Pleione and even the family Orchidaceae.

Disclosure statement
The authors declare that they have no competing interests.