The Utility of Graph Clustering of 5S Ribosomal DNA Homoeologs in Plant Allopolyploids, Homoploid Hybrids, and Cryptic Introgressants

Abstract

Introduction: Ribosomal DNA (rDNA) loci have been widely used for identification of allopolyploids and hybrids, although few of these studies employed high-throughput sequencing data. Here we use graph clustering implemented in the RepeatExplorer (RE) pipeline to analyze homoeologous 5S rDNA arrays at the genomic level searching for hybridogenic origin of species. Data were obtained from more than 80 plant species, including several well-defined allopolyploids and homoploid hybrids of different evolutionary ages and from widely dispersed taxonomic groups.Results: (i) Diploids show simple circular-shaped graphs of their 5S rDNA clusters. In contrast, most allopolyploids and other interspecific hybrids exhibit more complex graphs composed of two or more interconnected loops representing intergenic spacers (IGS). (ii) There was a relationship between graph complexity and locus numbers. (iii) The sequences and lengths of the 5S rDNA units reconstituted in silico from k-mers were congruent with those experimentally determined. (iv) Three-genomic comparative cluster analysis of reads from allopolyploids and progenitor diploids allowed identification of homoeologous 5S rRNA gene families even in relatively ancient (c. 1 Myr) Gossypium and Brachypodium allopolyploids which already exhibit uniparental partial loss of rDNA repeats. (v) Finally, species harboring introgressed genomes exhibit exceptionally complex graph structures. Conclusion:We found that the cluster graph shapes and graph parameters (k-mer coverage scores and connected component index) well-reflect the organization and intragenomic homogeneity of 5S rDNA repeats. We propose that the analysis of 5S rDNA cluster graphs computed by the RE pipeline together with the cytogenetic analysis might be a reliable approach for the determination of the hybrid or allopolyploid plant species parentage and may also be useful for detecting historical introgression events.

A chromosome-level reference genome of Ensete glaucum gives insight into diversity, chromosomal and repetitive sequence evolution in the Musaceae

“…The 5S rDNA locus lies in the middle of the short arm of chromosome eg05, in three rDNA arrays. Garcia et al [49] show the rather unusual and highly complex structure of 5S rDNA in M. acuminata using graph-based clusters of reads with multiple IGS and retroelement components, supporting the complexity reported here in the E. glaucum assembly. The multiple hybridization sites evident from the in situ hybridization site on one pair of chromosomes (Fig.…”

Ancient Origin of Two 5S rDNA Families Dominating in the Genus Rosa and Their Behavior in the Canina-Type Meiosis

“…Thus, in order to determine the number and genomic representation of individual 5S rRNA gene families, we applied clustering analysis ( Figure 3 ). The cluster graph shapes provide information about the number and type of 5S gene families revealing potential hybridization and introgression ( Garcia et al, 2020 ). It visualizes divergent IGS families as loops emanating from the bridge region which contains reads derived from a conserved coding region.…”

Personal Perspectives on Plant Ribosomal RNA Genes Research: From Precursor-rRNA to Molecular Evolution

“…Similarly, the 5S IGS appeared very useful for clarifying phylogenetic relationships at low taxonomic levels (Röser et al, 2001;Volkov et al, 2001;Denk and Grimm, 2010;Tynkevich and Volkov, 2019;Ishchenko et al, 2021) and for identification of interspecific hybrids (Garcia et al, 2020). This contrasts with the really strongly conserved sequences of the mature 5S, 5.8 S, 18S, and 25S rRNA coding regions.…”