The complete chloroplast genome sequence of Thunbergia erecta (Benth.) T. Anders. (Acanthaceae)

Abstract Thunbergia erecta (Benth.) T. Anders. is an upright shrub species of Acanthaceae with great ecological and economical values. In this paper, we explored the complete chloroplast genome sequence of T. erecta using next generation sequencing to provide genomic resources that could help to promote its conservation. The genome of T. erecta is 152,202 bp in length, containing a large single-copy region of 84,232 bp, and a small single-copy region of 17,656 bp. It encodes a total of 131 genes, including 8 rRNA genes, 37 tRNA genes, 84 protein-coding genes and 2 pseudo genes. The GC content of T. erecta genome is 38.47%. The phylogenetic analysis suggests that it was closely related to Avicennia marina in Acanthaceae. In addition, we found that T. erecta appeared late clade in the whole family while outgroups plants appeared even later. T. erecta is morphologically similar to other plants in Acanthaceae, but is genetically closed to the outgroup species.

Thunbergia erecta is a prominent species of Acanthaceae, around two meters tall, which is native to tropical West Africa and cultivated as ornamental plants around the world Tsui 2002, 2011). It possesses high value for medicinal (Refaey et al. 2021) and ornamental purposes and a good vertical greening material for scaffolds, flower fences and walls as well. However, little progress has been made to its complete chloroplast (cp) genome. In this work, we characterized the complete cp genome sequence of T. erecta, based on Illumina pair-end sequencing data, and deposited the sequence in GeneBank (MZ555773) to provide a valuable complete cp genome resource.
The fresh leaves were collected from Xishuangbanna Tropical Botanical Garden ( ÃÃ latitude ÃÃ 21.6840 and ÃÃ longitude ÃÃ 101.4677) in Jinghong, Yunnan, China. A specimen was deposited at the herbarium of Nanjing Forestry University (contact person: Xuehong Ma; E-mail: xue-hongma@njfu.edu.cn) under the voucher number NF2021038. According to the International Union for Conservation of Nature (IUCN) policy on endangered species research, the sample collection and the study was conducted with permission from Xishuangbanna Tropical Botanical Garden. The genomic DNA was extracted and then sequenced based on Illumina pair-end sequencing data. By applying ultrasound to break DNA, the fragments of DNA were passivated, repaired and bonded and selected by agarose gel electrophoresis. The sample of genome sequencing library was formed by PCR amplification, which was carried out on Illumina Novaseq platform with PE150 reads by Nanjing Genepioneer Biotechnologies Inc. (Nanjing, China).
The original reading was filtered by fastp (version 0.20.0), and the clean data were assembled into chloroplast genome using SPAdes (Bankevich et al. 2012). There were no uncertain bases in the assembly results. Then, the reference sequence (Genebank accession number: NC050991) was used for quality control after assembly, and the assembled genome was annotated using CpGAVAS2 (Shi et al. 2019). The complete cp genome sequences of species were acquired necessary from NCBI.
To reveal the phylogenetic evolution of T. erecta, we constructed a ML phylogenetic tree based on 16 cp genomes from Acanthaceae and 3 cp genomes as outgroups from 2 taxa (Pedaliaceae, Linderniaceae). The sequence aligment by MAFFT (Rozewicki et al. 2019), IQTREE (Garg and Biju 2021) was used to perform maximum Likelihood (ML) tree with the TVM þ FþR4 model. The bootstrap method was used to test the reliability of phylogeny with 1000 replicates.
The phylogenetic analysis suggests that T. erecta was closely related to Avicennia marina in Acanthaceae.
In addition, T. erecta was more likely appeared in late differentiation stage in the whole family succession while outgroups species did even later. T. erecta is morphologically similar to other species in Acanthaceae, but is genetically closed to those in outgroup. As the increasing of sample collection sequences of this taxa and the deepening of research, its status will become clearer.

Author contributions
Among the members of the author group, Lili Tong, Xiaogang Xu and Lu Tian contributed to substantial conception or design of the work; Lu Tian and Yao Cheng were in charge of acquisition, analysis, and interpretation of data for the work; Xiaogang Xu and Lili Tong, contributed to manuscript preparation (Drafting the work or revising it critically for important intellectual content); Lili Tong and Xiaogang Xu contributed to final approval of the version to be published. All authors agree on the final version and to be accountable for all aspects of the work.  Data availability statement