High-quality chromosome-level de novo assembly of the Trifolium repens

Abstract

Background White clover (Trifolium repens L.), an excellent perennial legume forage, is an allotetraploid native to southeastern Europe and southern Asia. It has high nutritional, ecological, genetic breeding, and medicinal values and exhibits excellent resistance to cold, drought, trample, and weed infestation. Thus, white clover is widely planted in Europe, America, and China; however, the lack of reference genome limits its breeding and cultivation. This study generated a white clover de novo genome assembly at the chromosomal level and annotated its components. Results The PacBio third-generation Hi-Fi assembly and sequencing methods generated a 1096 Mb genome size of T. repens, with contigs of N50 = 14 Mb, scaffolds of N50 = 65 Mb, and BUSCO value of 98.5%. The newly assembled genome has better continuity and integrity than the previously reported white clover reference genome; thus provides important resources for the molecular breeding and evolution of white clover and other forage. Additionally, we annotated 90,128 high-confidence gene models from the genome. White clover was closely related to Trifolium pratense and Trifolium medium but distantly related to Glycine max, Vigna radiata, Medicago truncatula, and Cicer arietinum. The expansion, contraction, and GO functional enrichment analysis of the gene families showed that T. repens gene families were associated with biological processes, molecular function, cellular components, and environmental resistance, which explained its excellent agronomic traits. Conclusions This study reports a high-quality de novo assembly of white clover genome obtained at the chromosomal level using PacBio Hi-Fi sequencing, a third-generation sequencing. The generated high-quality genome assembly of white clover provides a key basis for accelerating the research and molecular breeding of this important forage crop. The genome is also valuable for future studies on legume forage biology, evolution, and genome-wide mapping of quantitative trait loci associated with the relevant agronomic traits.

De novo genome assembly of white clover (Trifolium repens L.) reveals the role of Copy Number Variation in rapid environmental adaptation

“…Approximately 60% of the genome assembly was made up of repeated elements; half of these (30% of the genome) were retroelements, which closely matches retroelement proportions in the two recently published long-read assemblies [11,12]. The retroelements did not show any biased distribution between the two white clover subgenomes.…”

“…While native to Europe, white clover has become widely naturalized in mesic environments worldwide (from subtropical to boreal climates), and recent genomic research has linked its ability to become so widely adapted to its allopolyploid origin from two ecologically distinct diploid progenitor species [9]. White clover's importance both in agriculture and as an eco-evolutionary model system has spurred recent efforts to develop high-quality genetic and genomic resources for the species [9][10][11][12][13]. Most recently, two independent genome sequencing projects have utilized long-read sequencing technologies to overcome the scaffolding di culties associated with polyploid genome assembly [11,12].…”

Genome assembly of Melilotus officinalis provides a new reference genome for functional genomics

“…The data suggest that both sweet yellow clover and white clover in the self-comparisons had peaks at approximately 0.75 (Fig. 4 C) [ 37 , 38 ]. In addition, the WGD event occurred when the KS value of sweet yellow clover was 0.75 (Fig.…”

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

“…These assemblies will facilitate marker-assisted breeding programs for traits of agronomic importance and provide increased resolution and ability to identify the genomic basis of adaptation in this increasingly used model in evolutionary ecology and genetics. Together with an alternative and upcoming chromosome-level assembly ( Wang et al 2023 ) and other high-quality reference genomes in the genus ( Dluhošová et al 2018 ; Bickhart et al 2022 ; Shirasawa et al 2023 ), our haplotype-resolved assembly will be particularly useful for identifying structural variation and facilitate the development of pangenomic references ( Eizenga et al 2020 ) for which haplotype-resolved assemblies are an asset ( Garg et al 2022 ).…”