Haplotype-Resolved, Chromosome-Level Assembly of White Clover (Trifolium repens L., Fabaceae)

Abstract

White clover (Trifolium repens L.; Fabaceae) is an important forage and cover crop in agricultural pastures around the world and is increasingly used in evolutionary ecology and genetics to understand the genetic basis of adaptation. Historically, improvements in white clover breeding practices and assessments of genetic variation in nature have been hampered by a lack of high-quality genomic resources for this species, owing in part to its high heterozygosity and allotetraploid hybrid origin. Here, we use PacBio HiFi and chromosome conformation capture (Omni-C) technologies to generate a chromosome-level, haplotype-resolved genome assembly for white clover totaling 998 Mbp (scaffold N50 = 59.3 Mbp) and 1 Gbp (scaffold N50 = 58.6 Mbp) for haplotypes 1 and 2, respectively, with each haplotype arranged into 16 chromosomes (8 per subgenome). We additionally provide a functionally annotated haploid mapping assembly (968 Mbp, scaffold N50 = 59.9 Mbp), which drastically improves on the existing reference assembly in both contiguity and assembly accuracy. We annotated 78,174 protein-coding genes, resulting in protein BUSCO completeness scores of 99.6% and 99.3% against the embryophyta_odb10 and fabales_odb10 lineage datasets, respectively.

Keywords: allotetraploid; genome assembly; haplotype-resolved; legume; polyploidy.

Fig. 1

(A) Linkage map from the “DG” mapping population (Olsen et al. 2021, bottom, converted to physical positions in Mbp) with markers (vertical black lines in ideogram) connected to their physical positions in both the previous reference assembly (Griffiths et al. 2019, blue, upper left) and the current haploid assembly (green, upper right). Lines connecting markers to their physical position are colored red if they map to the wrong chromosome based on the linkage data or black if they map to the correct chromosome. (B) Circos plot of haploid mapping assembly consisting of 16 chromosomes. From outside to inside: chromosomes (green ideograms), GC%, gene density (red), repeat proportion (blue), and a photo of flowering Trifolium repens (credit: James Santangelo). GC%, gene density, and repeat density were estimated in 500 Kb windows with a 100 Kb step. (C) GENESPACE Riparian plot showing synteny between the T. occidentale (bottom) and T. pallescens (top) subgenomes of T. repens. Black lines show inferred translocations between nonhomoeologous chromosomes (N = 3), whereas white gaps within homoeologous chromosomes show inversions (N = 6; Chr_06 contains two nested inversions).