Characterization of the chloroplast genome of Calanthe henryi (Epidendroideae; Orchidaceae)

Abstract The first complete chloroplast (cp) genome sequences of an endemic and endangered species in China, Calanthe henryi, were reported in this study. The cp genome of C. henryi was 158,256 bp long, with two inverted repeat (IR) regions of 26,348 bp, a large single copy (LSC) region of 87,137 bp, and a small single copy (SSC) region of 18,423 bp. The cp genome of this species contained 113 genes, including 79 protein-coding genes, 4 ribosomal RNA genes, and 30 transfer RNA genes. The overall GC content was 36.7%. Phylogenetic analysis of 60 cp genomes within the subfamily of Epidendroideae suggests that C. henryi is closely related to C. bicolor.

Calanthe henryi Rolfe is a perennial plant in the subfamily of Epidendroideae (Orchidaceae) and usually grows in the evergreen forests from 1,600 m to 2,100 m. It is an endemic species in China and distributed in Hubei, Sichuan (Chen et al. 2009), Guizhou (Zhang 2007), Hunan (Yu et al. 2006) and Jiangxi (Xiao et al. 2017) provinces. At present, C. henryi has been regarded as the vulnerable endangered plant by IUCN (China Plant Specialist Group 2004). To promote the conservation of this species, we sequenced and analyzed the complete chloroplast (cp) genome of C. henryi using highthroughput sequencing technology.
The fresh leaf of C. henryi was collected from Yushe National Forest Park, Guizhou province, Southwest of China (N26 27 0 16 00 , E104 48 0 8 00 , 2,205 m). The specimen (lpssy0307) was deposited in the herbarium of the Liupanshui Normal University (LPSNU). Total genome DNA was extracted with CTAB method (Doyle and Doyle 1987), which was used for the library construction and sequencing on the Illumina HiSeq 2500 Platform. Approximately 2 Gb raw data were generated and used to de novo assemble the complete cp genome with SPAdes (Bankevich et al. 2012). All genes were annotated using PGA (Qu et al. 2019) with manual adjustment.
To determine the phylogenetic position of C. henryi within Epidendroideae, the complete cp genome sequences from 59 species of Epidendroideae and five species from Orchidoideae were downloaded from GenBank ( Figure 1). All the complete cp genome sequences were aligned using MAFFT version 7.0 (Katoh and Standley 2013). Phylogenomic tree was reconstructed with the maximum likelihood (ML) and Bayesian inference (BI) methods (Ronquist et al. 2012;Stamatakis 2014). The ML and BI analyses generated the same tree topology (Figure 1). As shown in the phylogenetic tree (Figure 1), the species of Calanthe formed one monophyletic clade and C. henryi was close to C. bicolor. The C. henryi cp genome reported in this study may provide useful resources for the development of ornamental and ecological value as well as robust phylogenetic study at deep level of Orchidaceae in the future.

Disclosure statement
No potential conflict of interest was reported by the author(s).

Data availability statement
The data that support the findings of this study are openly available in National Center for Biotechnology Information at https://www.ncbi.nlm. nih.gov/, reference number MT385870. Submissions are accessioned but the sequence record is not examined and deposited by the GenBank Figure 1. The maximum likelihood (ML) tree of Epidendroideae (Orchidaceae) inferred from the complete chloroplast genome sequences. ML bootstrap percentages (1,000 replicates) and Bayesian inference (BI) posterior probabilities are shown above clades if lower than 100 and 1.0 (dash indicate the brahches that are not supported by posterior probabilities).
annotation staff before it is free of errors or problems. Therefore, the live link to our dataset is not available at present.