Draft genome sequence of wild Prunus yedoensis reveals massive inter-specific hybridization between sympatric flowering cherries

Abstract

Background: Hybridization is an important evolutionary process that results in increased plant diversity. Flowering Prunus includes popular cherry species that are appreciated worldwide for their flowers. The ornamental characteristics were acquired both naturally and through artificially hybridizing species with heterozygous genomes. Therefore, the genome of hybrid flowering Prunus presents important challenges both in plant genomics and evolutionary biology.

Results: We use long reads to sequence and analyze the highly heterozygous genome of wild Prunus yedoensis. The genome assembly covers > 93% of the gene space; annotation identified 41,294 protein-coding genes. Comparative analysis of the genome with 16 accessions of six related taxa shows that 41% of the genes were assigned into the maternal or paternal state. This indicates that wild P. yedoensis is an F1 hybrid originating from a cross between maternal P. pendula f. ascendens and paternal P. jamasakura, and it can be clearly distinguished from its confusing taxon, Yoshino cherry. A focused analysis of the S-locus haplotypes of closely related taxa distributed in a sympatric natural habitat suggests that reduced restriction of inter-specific hybridization due to strong gametophytic self-incompatibility is likely to promote complex hybridization of wild Prunus species and the development of a hybrid swarm.

Conclusions: We report the draft genome assembly of a natural hybrid Prunus species using long-read sequencing and sequence phasing. Based on a comprehensive comparative genome analysis with related taxa, it appears that cross-species hybridization in sympatric habitats is an ongoing process that facilitates the diversification of flowering Prunus.

Keywords: Flowering Prunus; Hybrid genome; Long-read sequencing; S-locus haplotype; Sequence phase.

Fig. 1

The reference accession of wild Prunus yedoensis used in this study. a Photographs of a Pyn-Jeju2 tree and its flowers and berries taken from March to April 2017. b Estimation of the genome size of Pyn-Jeju2 based on K-mer analysis. The top panel represents the volume of K-17mer (Y-axis) plotted against the frequency at which it occurs (X-axis). The gray and black peaks correspond to heterozygous and homozygous reads, respectively. The bottom panel shows the estimated haploid genome size based on the homozygous K-mer peak as well as flow cytometry analysis

Fig. 2

Phasing and arrangement of the heterozygotic genome assembly. a Examples of haplotype-phased gene models. Gene models predicted from the initial “haplotype-fused” assembly are phased according to read mapping and SNP analysis using the Illumina short-read sequences of putative parental species. Genes were phased into one parental haplotype if a gene was aligned only by reads from one parental species (unique mapping) or had at least twofold as many supports for SNPs by reads of one parental species (phased by SNP). Genes with similar supports of read mapping for both parental species are defined as common type. Colored dots denote SNPs identified in the aligned reads. b Chromosomal arrangement of the gene-phased genome assembly of wild P. yedoensis (Pyn) onto the P. persica (Pp) genome. c Distribution of haplotype-phased genes in the tentative chromosomes of wild P. yedoensis. Colored dots or lines represent maternal-phased genes (red), paternal-phased genes (blue), or common genes (gray)

Fig. 3

Comparative analysis of the wild P. yedoensis var. nudiflora genome. a Venn diagram showing the unique and shared gene families between six sequenced genomes of the Rosaceae family. The number of gene families and genes (in bracket) for each group are shown. b Distribution of Ks values obtained from comparisons of orthologous gene sets between six genomes of the Rosaceae family and paralogous gene sets in wild P. yedoensis. c Genome evolution of Prunus species. Estimation of dates for speciation events are given in millions of years based on Bayesian evolutionary analysis of 276 conserved single copy genes. Pyn P. yedoensis var. nudiflora, Fv Fragaria vesca, Mt Medicago truncatula, Mxd Malus × domestica; Pa P. avium, Pm P. mume, Pp P. persica. Pal Paleocene, Eoc Eocene, Oli Oligocene, Mio Miocene, Pli Pliocene, Qua Quaternary

Fig. 4

Differential expression of haplotype-phased genes in various tissues of wild P. yedoensis. a Heat maps representing the expression of 562 maternal- and 576 paternal-phased genes, which are identified as differentially expressed genes in the mRNA-seq analysis, in different tissues. The normalized count values of a given gene from three independent biological replicates across all samples were used as a normalization factor. The vertical axes organize genes according to co-expression. The horizontal axes represent five tissues: leaf (L), petal (Pe), pistil (Pi), stamen (S), and berry (B). b Heat maps showing the differential expression of a selected category of genes related to development and secondary metabolite biosynthesis. The average normalized count values represent the relative expression across tissues. M maternal-phased genes, P paternal-phased genes

Fig. 5

The genomic relationships between flowering Prunus taxa. a Multidimensional scaling of Prunus accessions. Closely related accessions of Ppa (red square symbol), Pyn (green circle symbol), Pj or Psa (blue triangle symbol), and Pxy (black diamond symbol) are grouped together using dotted circles. b A maximum likelihood tree of Prunus accessions based on SNPs/InDels identified by variome analysis. The accession names are presented in Additional file 11: Table S10

Fig. 6

Characterization of S haplotypes in flowering Prunus species. a Microsynteny at the S-locus regions between wild P. yedoensis and fruit crop Prunus (P. persica and P. mume). There are two S haplotypes in the heterozygous Pyn genome (31 kb of S1 and 35 kb of S2) compared to a single S haplotype in the homozygous Pp and Pm genomes. Syntenic genes are connected with lines. b Relative expression levels of the S-RNase and SFB genes in different tissues are presented by the average fragments per kilobase million (FPKM) value from three independent biological replicates. c S haplotype network in a natural Prunus population. A total of 15 S haplotypes from 12 accessions, which are distributed sympatrically in a natural habitat on Jeju Island, were identified. Accessions are placed according to their relative geographic location in the natural habitat. Shared S haplotypes between accessions are connected with lines of the same color. Chloroplast genome lineage, showing < 10 nucleotide differences in the protein-coding sequences of the whole chloroplast DNA (Additional file 12: Table S11), is also presented in the green box