Biology subjects

Wagner, N. D.

Publications and source records attributed to Wagner, N. D..

8 recordsLinked to original sources

bioRxiv2023

Efficient hybrid strategies for assembling the plastome, mitochondriome, and large nuclear genome of diploid Ranunculus cassubicifolius (Ranunculaceae)

Currently, it is still a challenge - in terms of laboratory effort and cost, as well as assembly quality - to unravel the sequence of large and complex genomes from non-model plants. This often hampers the study of evolutionarily intricate species groups. The species-rich genus Ranunculus (Ranunculaceae) is an angiosperm model system for the study of polyploidy, apomixis, reticulate evolution, and biogeography. However, neither mitochondrial, nor high-quality nuclear genome sequences are available. This limits phylogenomic, functional, and taxonomic analyses thus far. Here, we tested Illumina short-read, Oxford Nanopore Technology (ONT) or PacBio/HiFi long-read, and hybrid-read assembly strategies. We used the diploid progenitor species R. cassubicifolius (R. auricomus complex), and selected the best assemblies in terms of completeness, contiguity, and quality scores. We first assembled the plastome (156 kbp, 85 genes) and mitogenome (1.18 Mbp, 40 genes) sequences using Illumina and Illumina-PacBio-hybrid strategies, respectively. We also present an updated plastome and the first mitogenome phylogeny of Ranunculaceae, including studies of gene loss (e.g., infA, ycf15, or rps) with evolutionary implications. For the nuclear genome, we favored a PacBio-based assembly three-times polished with filtered reads and subsequently scaffolded into 8 pseudochromosomes by chromatin conformation data (Hi-C) as the representative sequence. We obtained a haploid genome sequence with 2.69 Gbp, 94.5% complete BUSCO embryophyta_odb10 genes found, and 31,322 annotated genes. The genomic information presented here will improve phylogenomic analyses in this species complex, and will enable advanced functional, evolutionary, and biogeographic analyses for the genus and beyond Ranunculaceae in the future. Significance StatementThe genus Ranunculus is a model system in flowering plants for polyploidy, apomixis, evolution, and biogeography research. We present the first nuclear and mitochondrial genome sequence, and the plastid evolution of Ranunculaceae. Using Illumina, ONT, and PacBio data, we developed an efficient assembly strategy that can be applied to other non-model plants. Results presented here are useful for improving population genomic and phylogenomic analyses, and enable better functional analyses at species, genus and family level.

genomics↗

bioRxiv2021

Unraveling phylogenetic relationships, reticulate evolution, and genome composition of polyploid plant complexes by RAD-Seq and Hyb-Seq

Complex genome evolution of young polyploid complexes is poorly understood. Besides challenges caused by hybridization, polyploidization, and incomplete lineage sorting, bioinformatic analyses are often exacerbated by missing information on progenitors, ploidy, and reproduction modes. By using a comprehensive, self-developed bioinformatic pipeline integrating phylogenetic, structure, network, and SNP-origin analyses, we for the first time unraveled polyploid phylogenetic relationships and genome evolution within the large Eurasian Ranunculus auricomus species complex comprising more than 840 taxa. Our results rely on 97,312 genomic RAD-Seq loci, target enrichment of 576 nuclear genes (48 phased), and 71 plastid regions (Hyb-Seq; OMICS-data) derived from the 75 most widespread polyploid apomictic taxa and four di- and one tetraploid potential sexual progenitor species. Phylogenetic tree and structure analyses consistently showed 3-5 supported polyploid groups, each containing sexual progenitor species. In total, analyses revealed four diploid sexual progenitors and a one unknown, probably extinct progenitor, contributing to the genome composition of R. auricomus polyploids. Phylogenetic network, structure, and SNP-origin analyses based on RAD-Seq loci and phased nuclear genes completed by plastid data demonstrated predominantly allopolyploid origins, each involving 2-3 different diploid sexual subgenomes. Allotetraploid genomes were characterized by subgenome dominance and large proportions of interspecific, non-hybrid SNPs, indicating an enormous degree of post-origin evolution (i.e., Mendelian segregation of the diploid hybrid generations, back-crossings, and gene flow due to facultative sexuality of apomicts), but only low proportions of lineage-specific SNPs. The R. auricomus model system is the first large European polyploid species complex studied with reduced representation OMICS data. Our bioinformatic pipeline underlines the importance of combining different approaches and datasets to successfully unveil how reticulate evolution and post-origin processes shape the diversity of polyploid plant complexes.

evolutionary biology↗

bioRxiv2021

Using herbarium samples for NGS methods - a methodological comparison

Herbaria harbor a tremendous amount of plant specimens that are rarely used for plant systematic studies. The main reason is the difficulty to extract a decent quantity of good quality DNA from the preserved plant material. While the extraction of ancient DNA in animals is well established, studies including old plant material are still underrepresented. In our study we compared the standard Qiagen DNeasy Plant Mini Kit and a specific PTB-DTT protocol on two different plant genera (Xanthium L. and Salix L.). The included herbarium material covered about two centuries of plant collections. A selected subset of samples was used for a standard library preparation as well as a target enrichment approach. The results revealed that PTB-PTT resulted in higher quantity and quality regarding DNA yield. For relatively recent herbarium specimens, and despite the lower overall yield of DNA, the Qiagen Kit resulted in better sequencing results regarding the number of filtered and mapped reads. We were able to successfully sequence a sample from 1820 and conclude that it is possible to include old herbarium specimens in NGS approaches. This opens a treasure box for phylogenomic research.

plant biology↗

bioRxiv2020

Characterization of SARS-CoV-2 N protein reveals multiple functional consequences of the C-terminal domain

Nucleocapsid protein (N) is the most abundant viral protein encoded by SARS-CoV-2, the causative agent of COVID-19. N plays key roles at different steps in the replication cycle and is used as a serological marker of infection. Here we characterize the biochemical properties of SARS-CoV-2 N. We define the N domains important for oligomerization and RNA binding that are associated with spherical droplet formation and suggest that N accessibility and assembly may be regulated by phosphorylation. We also map the RNA binding interface using hydrogen-deuterium exchange mass spectrometry. Finally, we find that the N protein C-terminal domain is the most immunogenic by sensitivity, based upon antibody binding to COVID-19 patient samples from the US and Hong Kong. Together, these findings uncover domain-specific insights into the significance of SARS-CoV-2 N and highlight the diagnostic value of using N domains as highly specific and sensitive markers of COVID-19.

biochemistry↗

bioRxiv2020

Chromosome-scale assembly of the genome of Salix dunnii reveals a male-heterogametic sex determination system on chromosome 7

Sex determination systems in plants can involve either female or male heterogamety (ZW or XY, respectively). Here we used Illumina short reads, Oxford Nanopore Technologies (ONT) long reads, and Hi-C reads to assemble the first chromosome-scale genome of a female willow tree (Salix dunnii), and to predict genes using transcriptome sequences and available databases. The final genome sequence of 328 Mb in total was assembled in 29 contigs, and includes 31,501 genes. We inferred a male heterogametic sex determining factor on chromosome 7, suggesting that, unlike the female heterogamety of most species in the genus Salix, male heterogamety evolved in the subgenus Salix. The S. dunnii X-linked region occupies about 3.21 Mb of chromosome 7, and is probably in a pericentromeric region. Our data suggest that this region is enriched for transposable element insertions, and about one third of its 124 protein-coding genes were gained via duplications from other genome regions. We detect purifying selection on the genes that were ancestrally present in the region, though some have been lost. Transcriptome data from female and male individuals show more male- than female-biased genes in catkin and leaf tissues, and indicate enrichment for male-biased genes in the pseudo-autosomal regions. Our study provides valuable genomic resources for studying sex chromosome evolution in Salicaceae family.

genomics↗

bioRxiv2020

Conservation genomics of an Australian orchid complex with implications for the taxonomic and conservation status of Corybas dowlingii

This study assessed genomic diversity in an Australian species complex in the helmet orchids to clarify taxonomic delimitation and conservation status of the threatened species Corybas dowlingii, a narrow endemic from southeast Australia. Taxonomic delimitation between the three closely related species C. aconitiflorus, C. barbarae, and C. dowlingii has been mainly based on floral traits which exhibit varying degrees of overlap, rendering species delimitation in the complex difficult. Genomic data for the species complex was generated using double-digest restriction-site associated DNA (ddRAD) sequencing. Maximum likelihood, NeighborNet, and Bayesian structure analyses showed genetic differentiation within the species complex and retrieved genomic signatures consistent with hybridisation and introgression between C. aconitiflorus and C. barbarae, and an intermediate genetic position of C. dowlingii indicating a hybrid origin of the species. The genetic structure analysis showed varying levels of genetic admixture for several C. aconitiflorus, C. barbarae, and C. dowlingii samples, thus further corroborating the presence of hybridisation and introgression within the species complex. The taxonomic status of C. dowlingii D.L.Jones was revised to C. x dowlingii D.L.Jones stat. nov. to reflect its hybrid origin. The conservation status of C. x dowlingii was assessed based on key ecological and ethical aspects, and recommendations made regarding its conservation status in Australian conservation legislation.

genetics↗

bioRxiv2020

RAD sequencing data reveal a radiation of willow species (Salix L., Salicaceae) in the Hengduan Mountains and adjacent areas

The Hengduan Mountains (HDM) in South West China are an important hotspot of plant diversity and endemism and considered to be a secondary diversification center for the woody plant genus Salix (Salicaceae). This study aimed to reconstruct the spatio-temporal evolution of the Salix Chamaetia-Vetrix clade in the HDM and to test for the occurrence of a radiation. We inferred phylogenetic relationships based on more than 34,000 RAD loci of 27 species. Phylogenetic analyses recovered a well-resolved tree topology with two major clades, the Eurasian and the HDM clade and a divergence time of c. 23.9 Ma. The HDM clade comprises two subclades. The species of the HDM clade originated in north HDM and adjacent areas and then dispersed into the south HDM, westwards to the Himalayas and eastwards to the Qinling Mountains. Niche modelling analyses revealed that during the last glacial maximum, range contractions were observed in the northern areas, while southward expansions resulted in range overlaps. The reconstruction of putative adaptive character evolution of plant height, inflorescence and flower morphology indicate that adaptations to altitudinal distribution contributed to the diversification of the HDM willows. Our data indicate that a radiation occurred in HDM within the Salix Chamaetia-Vetrix clade. Dispersal within the mountain system and to adjacent regions as well as survival in glacial refugia have shaped the biogeographical history of the clade. Differentiation along altitudinal zonation concomitant to morphological adaptations to colder climates may be important ecological factors for the high species diversity of Salix in this area.

evolutionary biology↗

bioRxiv2019

Relationships and genome evolution of polyploid Salix species revealed by RAD sequencing data

AO_SCPLOWBSTRACTC_SCPLOWDespite the general progress in using next generation sequencing techniques for evolutionary research questions, the analysis of polyploid species is still hampered by the lack of suitable analytical tools and the statistical difficulties of dealing with more than two alleles per locus. Polyploidization and especially allopolyploidy leads to new combinations of traits by combining genomes of two or more parental species. This enhances the adaptive potential and often results in speciation. However, multiple origins of polyploids, backcrossing to the parental species and post-origin evolution can strongly influence the genome composition of polyploid species. Here, we used RAD sequencing, which revealed 23,393 loci and 320,010 high quality SNPs, to analyze the relationships and origin of seven polyploid species of the diverse genus Salix by utilizing a phylogenomic and a network approach, as well as analyzing the genetic structure and composition of the polyploid genome in comparison to putative parental species. We adapted the SNiPloid pipeline that was originally developed to analyse SNP composition of recently established allotetraploid crop lineages to RAD sequencing data by using concatenated RAD loci as reference. Our results revealed a well-resolved phylogeny of 35 species of Eurasian shrub willows (Salix subg. Chamaetia/Vetrix), including 28 diploid and 7 polyploid species. Polyploidization in willows appears to be predominantly connected to hybridization, i.e. to an allopolyploid origin of species. More ancient allopolyploidization events involving hybridization of more distantly related, ancestral lineages were observed for two hexaploid and one octoploid species. Our data suggested a more recent allopolyploid origin for the included tetraploids within the major subclades and identified putative parental taxa that appear to be plausible in the context of geographical, morphological and ecological patterns. SNiPloid and HyDe analyses disentangled the different genomic signatures resulting from hybrid origin, backcrossing, and secondary post-origin evolution in the polyploid species. All tetraploids showed a considerable post-origin, species-specific proportion of SNPs. The amount of extant hybridization appears to be related to the degree of geographical and ecological isolation of species. Our data demonstrate that high-quality RAD sequencing data are suitable and highly informative for the analysis of the origin and relationships of polyploid species. The combination of the traditional tools RAxML, STRUCTURE, SplitsTree and recently developed programs like SNAPP, HyDe and SNiPloid established a bioinformatic pipeline for unraveling the complexity of polyploid genomes.

evolutionary biology↗