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. 2024 Dec 3;9(12):1648–1652. doi: 10.1080/23802359.2024.2435901

The complete chloroplast genome sequence of Melothria scabra (Cucurbitaceae)

Chan Deng a,b, Xinbi Jia a,b, Siyue Chen a,b, Jiaqi Guo a,b, Chenghong Zeng a,b, Yuewen Chen a,b, Qianglong Zhu a,b,✉, Yingjin Huang a,b,✉
PMCID: PMC11619021  PMID: 39640867

Abstract

Melothria scabra has gradually become an economically important plant worldwide. The complete chloroplast genome of M. scabra has a length of 156,744 bp, contains a large single-copy (LSC) region (86,387 bp), a small single-copy (SSC) region (18,055 bp), and two inverted repeats (IRs) with the same length of 26,151 bp. In total, 126 genes were detected, including 83 protein-encoding genes, 35 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes. For phylogenetic analysis, M. scabra has a closer genetic relationship with Cucumis sativus and Citrullus lanatus. The complete chloroplast genome sequence of M. scabra would promote the germplasm exploration, phylogenetic relationships, and molecular biology researches in Melothria.

Keywords: Cucurbitaceae, chloroplast genome, Melothria scabra, phylogenetic analysis

Introduction

Melothria scabra is commonly referred to as the cucamelon, penguins, mini watermelon, or sour cucumber (Chomicki et al. 2020). M. scabra is a diploid (2n = 2x = 24) species that originated in Mexico and Central America; now, it has successfully spread to Asia, Europe, and Africa (Bhowmick & Jha 2022). M. scabra has angular grooves on its stem and branches, bright green leaves, palmate serrated leaves, small and pointed melon leaves, yellow flowers, and thin vines. It grows rapidly, has a strong covering ability, and is often used as an ornamental plant in gardens (Tao et al. 2013; Kamaruddin et al. 2021).

The appearance of M. scabra is similar to that of a mini watermelon, but its flavor is similar to that of a cucumber (Tao et al. 2013). The fruit flesh is rich in vitamin C, lycopene, calcium, iron, and several other essential nutrients, making it a new popular fruit (Dunaevskaya & Kravchenko 2020). Furthermore, the ethanol, methanol, and ethyl acetate fractions of M. scabra were shown to exhibit significant antioxidant activity, indicating that M. scabra could be a promising source of natural antioxidant agents (Kamaruddin et al. 2021). A preliminary investigation demonstrated that M. scabra could have antidiabetic and hypoglycemic properties (Govindula et al. 2019). In addition, relatively high concentrations of the amino acids arginine and citrulline, which have anticancer properties, were detected in M. scabra (Roberts et al. 2018).

However, M. scabra shares some interspecific morphological traits with other related species in the Cucurbitaceae family, such as watermelon and cucumber, which leads to a perplexing taxonomy for M. scabra. Furthermore, the lack of genetic diversity analyses of M. scabra is hindering the effective development and utilization of germplasm resources. The complete chloroplast genome is widely used in plant species taxonomy, phylogenetic, and genetic diversity analyses (Song et al. 2022; Zhang et al. 2022; Liu et al. 2024). Compared with nuclear genomes, the chloroplast genome is mostly maternally inherited, with independent evolutionary routes, moderate evolutionary rates, low nucleotide substitution rates, and stable gene structures, which can easily provide genetic information for plant species taxonomy studies (Kaundun & Matsumoto 2011). Therefore, we sequenced, assembled, and annotated the chloroplast genome of M. scabra for the first time in this study.

Materials and methods

Plant material, DNA extraction, and sequencing

M. scabra (voucher number: QZ01MS, Figure 1) seeds were collected from a cultivated population in October 2023 in Feidong County (N31°93′12.81″, E117°63′96.69″), Anhui Province, China. They were subsequently planted in the horticulture teaching practice base at Jiangxi Agricultural University, Nanchang, China (contact person: Chan Deng, dengchan2023@163.com). During the healthy growth of plants, healthy and fresh leaves were selected in November 2023, and their total genomic DNA was extracted via a modified cetyltrimethylammonium bromide (CTAB) method (Li et al. 2013). The concentration and quality of the DNA samples were detected via agarose gel electrophoresis and a Nanodrop-2000 nucleic acid assay, and the DNA samples that met the requirements were used for subsequent experiments. Subsequently, 20 μg of isolated gDNA was sent to Bena Technology Co. Ltd. for genome sequencing via the Illumina NovaSeq 6000 platform (Wuhan, China).

Figure 1.

Figure 1.

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The growth picture of M. scabra (3520 × 2640). The image was taken in june 2024 by Chan Deng in the horticulture teaching practice base, at Jiangxi Agricultural University, Nanchang, China (N28°45′51″, E115°49′58″). M. scabra have bright green and small leaves, curly vines, and yellow flowers. Meanwhile, the fruit is soval with an average length of 2.4 cm and a width of 1.5 cm.

Chloroplast genome assembly and annotation

The raw pair-end (PE) reads were filtered for quality control to ensure the high quality of the clean PE reads via NGSQCToolkit v2.3.3. The high-quality clean PE reads were randomly selected via Seqtk, and then the clean PE reads were assembled into contig sequences via the Plasmidspades.py script program in SPAdes v3.6.1 (Bankevich et al. 2012). The scaffolds were subsequently aligned to the chloroplast reference genome of Citrullus lanatus (NC_032008.1) via BLASTN (Talamantes et al. 2021), and the draft chloroplast genome of M. scabra was constructed.

The gaps in the draft chloroplast genome of M. scabra were filled via GapCloser. The complete chloroplast genome sequence was subsequently annotated via CPGAVAS2 and GeSeq (Tillich et al. 2017; Shi et al. 2019), and the annotation results were subsequently checked and corrected via Sequin. A circular map of the complete chloroplast genome of M. scabra was constructed via CPGView (Liu et al. 2023).

Phylogenetic analysis

To clarify the phylogenetic relationships among M. scabra and other related species in Cucurbitaceae in this study, the complete chloroplast genome sequences were aligned via MAFFT, v7.463 (Rozewicki et al. 2019). The phylogenetic tree was constructed via MEGA v11 (Tamura et al. 2021) with the maximum likelihood (ML) method and 1000 bootstrap values.

Results

General features of the chloroplast genome

A 0.4 Gb of high-quality clean PE reads was extracted from total sequence data for chloroplast genome assembly. The complete chloroplast genome of M. scabra is 156,744 bp in length and has a typical quadripartite structure (Figure 2A). The assembled complete chloroplast genome presented an average read mapping depth of 4174.60 × and a minimum mapping depth of 67 × (Figure S1). It contains LSC (86,387 bp) and SSC (18,055 bp) regions and two IRs (26,151 bp), and its total GC content is 37.19%. Furthermore, a total of 126 genes were annotated, including 83 protein-coding genes, 35 tRNA genes and eight rRNA genes. Among these genes, eleven (ndhB, rpl23, rps7, rrn16S, rrn4.5S, rrn5S, trnE-UUC, trnL-CAA, trnN-GUU, trnV-GAC, ycf2) are duplicates, six protein-coding genes (rps16, atpF, rpoC1, rpl2, ndhB, ndhA) have a single intron, and two protein-coding genes (ycf3, clpP) have two introns (Figure 2B and 2C). The annotated complete chloroplast genome of M. scabra was submitted to the sequence database of CNGBdb with the accession number N_001486263.

Figure 2.

Figure 2.

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The complete chloroplast genome map of M. scabra. (A) The genes are arranged on the circular chloroplast genome with different colored boxes representing specific gene regions. The genes transcribed in clockwise and counterclockwise directions are shown on the inner and outer sides of the circle, respectively. Additionally, the GC content is presented via the grey area in the center of the circle, while the four-part structures of LSC, SSC, IRa, and IRb are depicted on the inner circle. (B) The black-white arrow showed the cis-spliced genes, and (C) The black-grey-white arrow showed trans-spliced genes.

Phylogenetic relationship analysis

To date, the complete chloroplast genome of M. scabra is the first to be published in the genus Melothria, with 13 species. More than 60 complete chloroplast genomes belonging to 24 genera in the family Cucurbitaceae have been published in the NCBI genome database. To confirm the phylogenetic status of M. scabra, 25 complete chloroplast genome sequences representing the different genera of Cucurbitaceae and an outgroup species (Vitis vinifera) were used to infer a phylogenetic tree. A phylogenetic tree was constructed to explore the phylogenetic relationships between M. scabra and other Cucurbitaceae plants. The bootstrap value for the clade of M. scabra with other close relatives was 100, indicating that the phylogenetic tree has high reliability. The inferred phylogenetic tree revealed that M. scabra fell between Cucurbita and Cucumis, suggesting that M. scabra is a member of the family Cucurbitaceae. Moreover, M. scabra is closely related to Cucumis sativus and Citrullus lanatus, confirming that M. scabra shares interspecific morphological traits with Cucumis sativus and Citrullus lanatus (Figure 3).

Figure 3.

Figure 3.

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The phylogenetic tree suggested the relationship between M. scabra and 25 other species within the family Cucurbitaceae. The Vitis vinifera L. was taken as an outgroup. The maximum-likelihood method was used to infer the position of the M. scabra in the phylogenetic tree, using 1000 as the bootstrap value and the number of upper branches as the bootstrap value. The following GenBank sequences were used: Ampelosycios Hublot NC_046869.1, Bryonia marmorata NC_046865.1, Baijiania yunnanensis NC_046871.1, Benincasa hispida NC_056352.1, Citrullus lanatus NC_032008.1 (Zhu et al. 2016), Cucumis sativus NC_007144.1 (Plader et al. 2007), Cucurbita maxima NC_036505.1, Cyclanthera pedata NC_046860.1, Cyclantheropsis parviflora NC_046870.1, Cionosicys macranthus NC_046861.1, dendrosicyos socotranus NC_046862.1 (Bellot et al. 2020), Gynostemma Cardiospermum NC_035959.1, Herpetospermum pedunculosum NC_046858.1, Hodgsonia heteroclita NC_046857.1, Hemsleya lijiangensis NC_039653.1 (Zhang et al. 2018), Indofevillea khasiana NC_046859.1, Lagenaria siceraria NC_036808.1, Momordica sessilifolia NC_046872.1, Momordica charantia NC_036807.1, Nothoalsomitra suberosa NC_046876.1, Siraitia grosvenorii NC_043881, Sicyos edule NC_051498.1 (Cui et al. 2021), Trichosanthes baviensis NC_046864.1, Thladiantha dubia NC_046855.1, Vitis vinifera NC_081805.1 (Wang et al. 2021).

Discussion and conclusion

In this study, the complete chloroplast genome of M. scabra was sequenced, assembled de novo, annotated, and phylogenetic analysis was conducted with another related species in the Cucurbitaceae family. The chloroplast genome of M. scabra is 156,744 bp in size with a typical circular tetrameric structure and contains LSC and SSC regions and two IR regions. All 126 genes were identified, including 83 protein-encoding genes, 35 tRNA genes, and eight rRNA genes. There was no marked difference in the chloroplast genome structure or gene content between M. scabra and other related species in the Cucurbitaceae (Li et al. 2022; Zhou et al. 2023). Moreover, phylogenetic relationship analysis revealed that M. scabra belongs to the family Cucurbitaceae, which confirmed the phylogenetic relationships of M. scabra proposed in earlier studies (Kocyan et al. 2007; Chomicki et al. 2020). Furthermore, M. scabra and three other species (Cucumis sativus, Lagenaria siceraria, and Citrullus lanatus) were grouped into a clade, suggesting that the four species have relatively close genetic distances and that M. scabra could be the closest wild ancestor of the other three species. This study also supported the early conclusion that M. scabra looks like a mini watermelon and has a cucumber-like flavor (Tao et al. 2013). Our research results could be used to analyze the genetic diversity and phylogenetic relationships of M. scabra and its related species within the family Cucurbitaceae.

Supplementary Material

supplementary material.docx
TMDN_A_2435901_SM4903.docx (110KB, docx)

Funding Statement

This work was supported by grants from the Postdoctoral Research Funding program of Jiangxi Province (2021KY38)].

Ethical approval

No permits were required for the sample acquisition of Melothria scabra, and the study did not include any endangered or protected species.

Author contributions

C. Deng and X. B. Jia analyzed the sequence data and wrote the manuscript; S. Y. Chen and J. Q. Guo grew a plant and extracted a DNA sample, Y. W. Chen and C. H. Zeng performed the experiment data collection; Q. L. Zhu and Y. J. Huang designed the funded the study.

Disclosure statement

The authors declare no potential conflict of interest.

Data availability statement

The data supporting the findings of this study is available in CNGBdb (https://db.cngb.org) with the accession number: N_001486263. The associated project, sample, and experiment numbers are CNP0005296, CNS1008401, and CNX0953946 in the CNGBdb database, respectively.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

supplementary material.docx
TMDN_A_2435901_SM4903.docx (110KB, docx)

Data Availability Statement

The data supporting the findings of this study is available in CNGBdb (https://db.cngb.org) with the accession number: N_001486263. The associated project, sample, and experiment numbers are CNP0005296, CNS1008401, and CNX0953946 in the CNGBdb database, respectively.

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