The complete chloroplast genome of a new candidate cultivar, Sang Jae, of Abeliophyllum distichum Nakai (Oleaceae): initial step of A. distichum intraspecies variations atlas.

Chloroplast genome of Sang Jae, new candidate cultivar of Abeliophyllum distichum Nakai, has been sequenced to reveal infraspecific relationship of wildtypes and cultivars. Its length is 156,008 bp long (GC ratio is 37.8%) and has four subregions: 86,773 bp of large single copy (35.8%) and 17,827 bp of small single copy (31.9%) regions are separated by 25,704 bp of inverted repeat (43.2%) regions including 133 genes (87 protein-coding genes, eight rRNAs, and 37 tRNAs). Phylogenetic trees show that A. distichum is clustered with twelve Forsythia species and intraspecies variations level is similar to Olea europaea.

Abeliophyllum distichum Nakai, is endemic to Korea and monotypic species of genus Abeliophyllum (Nakai 1919; Melchior 1964). A novel candidate cultivar of A. distichum, Sang Jae, has been discovered, which has remarkable phenotypic differences from wildtype: flower is yellow and curved petal has wavy margin (Figure 1(A)) and fruit shape is obovate, different from that of wild type (Figure 1(B)). A. distichum is widely used as landscaping tree even though its natural habitats are limited and threatening (Kim and Maunder 1998; Lim et al. 2009; Lee et al. 2014). Four chloroplast genomes of two A. distichum are available presenting intraspecies relationship between cultivars and natural isolates (Kim et al. 2016; Ha et al. 2018; Park, Kim et al. 2019; Park, Min et al. 2019). Here, we completed third cultivar chloroplast genome of A. distichum for understanding intraspecies variations.

Figure 1.

(A) Pictures of flowers of Sang Jae and wildtype of A. distichum. (B) Pictures of fruits of Sang Jae and wildtype of A. distichum. (C) Maximum likelihood (bootstrap repeat is 1,000) and neighbor joining (bootstrap repeat is 10,000) phylogenetic trees of forty-eight Oleaceae chloroplast genomes: Abeliophyllum distichum (MN127986 in this study, MN116559, MK616470, MF407183, and NC_031445), Forsythia x intermedia (NC_036982), Forsythia giraldiana (MF407174), Forsythia likiangensis (MF407177), Forsythia suspensa (NC_036367, MF407180), Forsythia europaea (MF407184), Forsythia japonica (MF407175), Forsythia velutina (MF407181), Forsythia ovata (MF407178), Forsythia saxatilis (MF407179), Forsythia koreana (MF407176), Forsythia viridissima (MF407182), Fontanesia phillyreoides subsp. fortunei (MG255754), Olea woodiana subsp. woodiana (NC_015608), Olea exasperata voucher A. Costa 1 (NC_036985), Olea europaea subsp. maroccana (NC_015623), Olea europaea subsp. guanchica isolate La Gomera 10 (MG255764), Olea europaea cultivar Frantoio (GU931818), Olea europaea cultivar Bianchera (NC_013707), Olea europaea subsp. europaea cultivar Manzanilla (FN996972), Olea europaea subsp. europaea isolate Stavrovouni 11 (HF558645), Olea europaea subsp. laperrinei isolate Adjelella 10 (MG255765), Olea europaea subsp. europaea isolate Vallee du Fango 5 (MG255762), Olea europaea subsp. cuspidate (NC_015604), Hesperelaea palmeri (NC_025787), Chionanthus retusus (NC_035000), Olea europaea subsp. europaea isolate Oeiras 1 (MG255763), Schrebera arborea (NC_036986), Fraxinus excelsior (NC_037446), Syringa vulgaris (NC_036987), Noronhia lowryi (NC_036984), Fraxinus chiisanensis (MF980720), Chionanthus rupicola (NC_036980), Olea europaea subsp. cuspidate (FN996944), Olea europaea subsp. cuspidata isolate Menagesha Forest 14 (MG255760), Olea europaea subsp. cuspidata isolate Almihwit 5.1 (FN996943), Chionanthus parkinsonii (NC_036979), Syringa pinnatifolia (MG917095), Forestiera isabelae (NC_036981), Olea europaea subsp. europaea isolate Stavrovouni Monastery 11 (MG255761), Nestegis apetala (NC_036983), Ligustrum lucidum (MH394207), and Jasminum nudiflorum (NC_008407). Phylogenetic tree was drawn based on neighbor joining tree. The numbers above branches indicate bootstrap support values of maximum likelihood and neighbor joining phylogenetic trees, respectively.

Total DNA of Sang Jae collected in GoesanBunjae-Nongwon (Goesan-gun, Chungbuk Province, Republic of Korea, Voucher in InfoBoss Cyber Herbarium (IN); Y. Kim, IB-01026) was extracted from fresh leaves by using a DNeasy Plant Mini Kit (QIAGEN, Hilden, Germany). Genome was sequenced using HiSeqX at Macrogen Inc., Korea, and de novo assembly and sequence confirmation were done by Velvet 1.2.10 (Zerbino and Birney 2008), SOAPGapCloser 1.12 (Zhao et al. 2011), BWA 0.7.17 (Li 2013), and SAMtools 1.9 (Li et al. 2009). Geneious R11 11.0.5 (Biomatters Ltd., Auckland, New Zealand) was used for chloroplast genome annotation based on A. distichum chloroplast (MK616470; Park et al. 2019).

Chloroplast genome of A. distichum, Sang Jae, (Genbank accession is MN127986) is 156,008 bp (GC ratio is 37.8%) and has four subregions: 86,773 bp of large single copy (35.8%) and 17,827 bp of small single copy (31.9%) regions are separated by 25,704 bp of inverted repeat (IR; 43.2%). It contains 133 genes (87 protein-coding genes, eight rRNAs, and 37 tRNAs); 19 genes (7 protein-coding genes, 4 rRNAs, and 7 tRNAs) are duplicated in IR regions. It is identical to that of Ok-Hwang-1-ho because this cultivar was rescued from the same region.

Based on alignments with four A. distichum chloroplast genome, 9 single nucleotide polymorphisms (SNPs) and 11 insertions and deletions (INDELs) were found between Sang Jae and Dae Ryun (MN116559). While there are 93 SNPs and 56 INDELs between Sang Jae and natural isolate (NC_031445) caused by exceptional sequence variations (56 SNPs and 15 INDELs) in rpoC2. All INDELs do not affect frame of rpoC2. Only one INDEL was found in the comparison of another natural isolate (MF407183).

Forty-eight Oleaceae chloroplast genomes including five A. distichum chloroplast genomes were used for constructing bootstrapped neighbor joining and maximum likelihood phylogenic trees using MEGA X (Kumar et al. 2018) via modifying directions of subregions and aligning whole chloroplast genomes by MAFFT 7.388 (Katoh and Standley 2013). Phylogenetic trees show that five A. distichum form an independent clade from Forsythia species, supporting that genus Abeliophyllum has evolved separated from Forsythia clade. In addition, intraspecies variations among five A. distichum is similar to those of Olea europaea.