The complete chloroplast genome of a centennial olive tree (Olea europaea, Oleaceae) from the southern Peruvian coast

Introduction

The olive Olea europaea (Oleaceae) is a subtropical tree distributed on most continents in tropical and temperate environments (Dupin et al., 2020). The genus Olea comprises about 40 old world species (Jensen et al., 2002) and the majority of Oleaceae species possess an economic importance as most olive fruits are used for oil extraction. Vegetable oil is important for its nutritional and healthy advantages compared to others (Rallo et al., 2000). Currently, ten olive tree chloroplast (cp) genomes were reported from Mediterranean region (Besnard et al., 2011), Italy, Spain, China (Niu et al., 2020), and Jordan (Haddad et al., 2021). However, to date, the olive tree chloroplast genome from Peru has not been reported. In this study, we sequenced, assembled, and annotated for the first time the complete chloroplast genome of a centennial the olive tree from Peru ( Figure 1), using next generation sequencing (NGS), providing valuable information for genetic and evolutionary studies in the genus Olea.

Figure 1. Picture of Olea europaea, a centennial olive tree from Moquegua, Peru.

Methods

Plant materials and extraction of genomic DNA Fresh young leaves were collected from “El Algarrobal” district located in the Ilo province from Moquegua region (latitude 17.61294, longitude -71.27136). The specimen (MOQ001169) was deposited at the National University of Moquegua Herbarium (http://www.hmoqueguensis.unam.edu.pe/, Hibert Huaylla-Limachi hmoqueguensis@unam.edu.pe) under the voucher Nro. 001169. DNA was extracted using CTAB method (Doyle and Doyle 1990) with minor modifications for this specie. The quality and the quantity were evaluated on a 1% agarose gel and fluorescence using the Qubit™ 4 Fluorometer (Invitrogen, Waltham, MA, USA), respectively.

DNA sequence and genome assembly Library construction pair-end reads and sequencing were carried out using Illumina HiSeq 2500 platform and a PE 150 library, with the NexteraXT DNA Library Preparation Kit (Illumina, San Diego, CA, USA). We removed the adapters and verified the quality of reads employing Trim Galore (Martin, 2011) with arguments: −F embplant_pt −R 15 –reduce-reads-for-coverage inf. The cp genome was assembled with GetOrganelle v1.7.2 (Jin et al., 2020), using Olea europaea subsp. europaea (NC_015401) as reference. SPAdes v3.11.1 (Bankevich et al., 2012), bowtie2 v2.4.2 (Langmead and Salzberg, 2012), and BLAST+ v2.11 (Camacho et al., 2009) were also used in the pipeline with default settings.

Results

Olea europaea chloroplast genome assembly

The complete chloroplast genome of olive was 155,886 bp in length with typical quadripartite structure that included a large single-copy (LSC) of 86,610 bp and a small single copy region (SSC) of 17,790 bp separated by two inverted repeat (IR) regions (25,741 bp). Following of annotation and subsequent modifications, we submitted the complete chloroplast (cp) genome sequence to the GenBank database. This submission is associated with the accession number: ON767107. The average depth of coverage is 17,673.05X (Supplementary Figure 1, https://doi.org/10.5281/zenodo.14061216). The chloroplast genome contains 124 genes that consisted of 80 protein-coding genes, 36 tRNA, 8 rRNA. Of the total genes reported 12 present one intron. PafI and clpP1 genes contained two introns. We further report that most genes are present as a single copy, except 14 genes that were duplicated in IR regions ( Figure 2). A total of 11 cis-splicing genes (rps16, atpF, rpoC1, pafl, clpP1, petB, petD, rpl16, rpl2, ndhB, ndhA) were identified; one trans-splicing gene (rps12) was also identified (Supplementary Figure 2, Supplementary Figure 3).

Figure 2. Schematic map of features of Olea europaea chloroplast genome (Genes drawn outside the outer circle are transcribed clockwise, and those inside are transcribed counter-clockwise.

Genes belonging to different functional groups are color-coded. The genes functions are indicated colored in the bottom left corner. The dark grey inner circle indicates the presence of nodes in the LSC, SSC, IR regions).

Phylogenetic inference of Olea europaea

This study examined 19 species of the Oleaceae family, with one outgroup, Calycophyllum spruceanum (OK326865), to understand their evolutionary relationships by analyzing complete chloroplast genome sequences. Maximum likelihood (ML) analysis revealed three distinct monophyletic groups within the tribes Jasminenae, Oleeae, and Forsythiae, with 17 of the nodes showing 100% bootstrap support. In addition, O. europaea is shown to be sister to O. europaea subsp. Maroccana, and sister to them is O. europaea subsp. cuspidata ( Figure 3). As expected, Olea europaea was categorized in the Oleeae tribe. These phylogenetic trees are consistent with the established classification of the Oleaceae family.

Figure 3. The phylogenetic position for O. europaea according to the ML phylogenetic tree constructed based on 19 chloroplast genomes: Jasminum tortuosum NC_034691 (Unpublished), Jasminum sambac NC_034694 (Unpublished), Jasminum fluminense NC_042272 (Olofsson et al., 2019), Jasminum nudiflorum NC_008407 (Lee et al., 2007), Jasminun polyanthum NC_ 042273 (Olofsson et al., 2019), Olea europaea subsp. maroccana NC_015623 (Besnard et al., 2011), Olea europaea subsp. cuspidata NC_015604 (Besnard et al., 2011), Olea lancea NC_042278 (Olofsson et al., 2019), Olea paniculata NC_042460 (Olofsson et al., 2019), Osmanthus aff. armatus Besnard NC_042263 (Olofsson et al., 2019), Osmanthus cooperi NC_053565 (Wang et al., 2019), Picconia excelsa NC_042466 (Olofsson et al., 2019), Forestiera angustifolia NC_042782 (Olofsson et al., 2019), Forestiera isabelae NC_036981 (Van de Paer et al., 2018), Forsythia mandschurica NC_048504 (Olofsson et al., 2019), Forsythia x intermedia NC_036982 (Park et al., 2019), Abeliophyllum distichum NC_031445 (Kim et al., 2016), Myxopyrum hainanense NC_047485 (Zhu et al., 2020), Calycophyllum spruceanum OK326865 (Saldaña et al., 2022).

Discussion and conclusions

In this study, we assembled for the first time the chloroplast genome sequence of the centennial olive tree (Olea europaea) from the southern Peruvian coast. The results differ from other chloroplast genomes of Olea europaea, such as O. europaea subsp. europaea (NC_015401), which presents 85 protein-coding genes, 37 tRNA, and 8 rRNA (Haddad et al., 2021). Niu et al. (2020) reported that two varieties of O. europaea, subsp. europaea var. sylvestris and subsp. cuspidata, presented 133 genes, including 87 protein-coding genes, 37 tRNA, and 9 rRNA. Likewise, they found that the chloroplast genome length for both species were 155,886 bp (Niu et al., 2020). The phylogenetic results showed O. europaea is sister to O. europaea subsp. maroccana and O. europaea subsp. cuspidata in independent sister clade to other species of genus Olea the chloroplast genome sequence and annotation were submitted to NCBI with accession number ON767107.1.

Peruvian olive chloroplast genome will stimulate additional work to develop molecular markers for their application in a genomic selection program of superior individuals in early stages. In addition, this study may be a basis to know the location of genes of interest that may be used in gene editing. This work promotes knowing more about the genetics of the Peruvian olive tree, promoting its modern genetic improvement and conservation.

Authors’ contribution

C.L.S. analyzed the data, was also involved in drafting the article. L.C.-L., C.I.A. and R.E. analyzed the data. E.F.-H., F.Z.-V., J.C.G.-A., C.A.-G., D.L.G.-R. were involved in the conception and design of the work. L.C.-L. and F.Z.-V. were involved in sample collection. F.Z.-V., J.C.G.-A., P.I. and C.I.A. were involved in data interpretation.

F.Z.-V., J.C.G.-A., P.I. and C.I.A. were involved in funding acquisition. C.I.A was involved in drafting the article. All authors read and approved the manuscript.

Ethical approval

Olive sample collected and employed in this work do not involve protection determined by the Republic of Peru. Consequently, this study was exempted from ethical approval.