Biology subjects

Koelliker, R.

Publications and source records attributed to Koelliker, R..

7 recordsLinked to original sources

bioRxiv2023

High-resolution bulked segregant analysis enables candidate gene identification for bacterial wilt resistance in Italian ryegrass

Bacterial wilt, caused by Xanthomonas translucens pv. graminis (Xtg), is a serious disease of economically important forage grasses, including Italian ryegrass (Lolium multiflorum Lam.). A major QTL for resistance to Xtg was previously identified, but the precise location as well as the genetic factors underlying the resistance are yet to be determined. To this end, we applied a bulked segregant analysis (BSA) approach, using whole-genome deep sequencing of pools of the most resistant and most susceptible individuals of a large (n = 7,484) biparental F2 population segregating for resistance to Xtg. Using chromosome-level genome sequences as references, we were able to define a [~]300 kb region highly associated to resistance on pseudo-chromosome 4. Further investigation of this region revealed multiple genes with a known role in disease resistance, including genes encoding for Pik2-like disease resistance proteins, cysteine-rich kinases, and RGA4- and RGA5-like disease resistance proteins. Investigation of allele frequencies in the pools and comparative genome analysis in the grandparents of the F2 population revealed that some of these genes contain variants with allele frequencies that correspond to the expected heterozygosity in the resistant grandparent. This study emphasizes the efficacy combining BSA studies in very large populations with whole genome deep sequencing and high-quality genome sequences to pinpoint regions associated with a binary trait of interest and accurately define a small set of candidate genes. Furthermore, markers identified in this region hold significant potential for marker-assisted breeding strategies to breed resistance to Xtg in Italian ryegrass cultivars more efficiently.

plant biology↗

bioRxiv2023

An improved chromosome-level genome assembly of perennial ryegrass (Lolium perenne L.)

This work is an update and extension of the previously published article "Ultralong Oxford Nanopore Reads Enable the Development of a Reference-Grade Perennial Ryegrass Genome Assembly", by Frei et al.. The published genome assembly of the doubled haploid perennial ryegrass (Lolium perenne L.) genotype Kyuss marked a milestone for forage grass research and breeding. However, order and orientation errors may exist in the pseudo-chromosomes of Kyuss, since barley (Hordeum vulgare L.), which diverged 30 million years ago from perennial ryegrass, was used as the reference to scaffold Kyuss. To correct for structural errors possibly present in the published Kyuss assembly, we de novo assembled the genome again and generated 50-fold coverage high-throughput chromosome conformation capture (Hi-C) data to assist pseudo-chromosome construction. The resulting new chromosome-level assembly showed improved quality with high contiguity (contig N50 = 120 Mb), high completeness (total BUSCO score = 99%), high base-level accuracy (QV = 50) and correct pseudo-chromosome structure (validated by Hi-C contact map). This new assembly will serve as a better reference genome for Lolium spp. and greatly benefit the forage and turf grass research community.

genomics↗

bioRxiv2023

Higher seed yield through selection for reduced seed shattering in Italian ryegrass (Lolium multiflorum Lam.)

Seed shattering, i.e., the loss of seeds at ripening stage shortly before or during seed harvest, is strongly reducing seed yield in Italian ryegrass (Lolium multiflorum Lam.). The aim of this study was to evaluate the possibility to reduce seed shattering within breeding germplasm via recurrent phenotypic selection on spaced plants. Starting from a founder population of 300 plants serving as F0 population, two cycles of phenotypic selection for high and low seed shattering were applied and compared to randomly selected individuals on spaced plant level and in plot trials. Comparison of the five resulting populations in a spaced plant trial revealed a significant effect of selection, with lowest seed shattering (15.3%) observed in the population selected twice for decreased shattering (15.3%) and highest seed shattering (47.9%) for the population selected twice for increased shattering. The same ranking of the five F2 populations was observed in a subsequent trial with sown plots. Thus, using the method presented here, recurrent selection on single spaced plants allows to efficiently reduce seed shattering and, therefore, to increase seed yield in swards.

plant biology↗

bioRxiv2023

High genomic plasticity and unique features of Xanthomonas translucens pv. graminis revealed through comparative analysis of complete genome sequences

BackgroundXanthomonas translucens pv. graminis (Xtg) is a major bacterial pathogen of economically important forage grasses, causing severe yield losses. So far, genomic resources for this pathovar consisted mostly of draft genome sequences, and only one complete genome sequence was available, preventing comprehensive comparative genomic analyses. Such comparative analyses are essential in understanding the mechanisms involved in the virulence of pathogens and to identify virulence factors involved in pathogenicity. ResultsIn this study, we produced high-quality, complete genome sequences of four strains of Xtg, complementing the recently obtained complete genome sequence of the Xtg pathotype strain. These genomic resources allowed for a comprehensive comparative analysis, which revealed a high genomic plasticity with many chromosomal rearrangements, although the strains were highly related, with 99.9 to 100% average nucleotide identity. A high number of transposases were exclusively found in Xtg and corresponded to 413 to 457 insertion/excision transposable elements per strain. These mobile genetic elements are likely to be involved in the observed genomic plasticity and may play an important role in the adaptation of Xtg. The pathovar was found to lack a type IV secretion system, and it possessed the smallest set of type III effectors in the species. However, three XopE and XopX family effectors were found, while in the other pathovars of the species two or less were present. Additional genes that were specific to the pathovar were identified, including a unique set of minor pilins of the type IV pilus, 17 TonB-dependent receptors (TBDRs), and 11 degradative enzymes. ConclusionThese results suggest a high adaptability of Xtg, conferred by the abundance of mobile genetic elements, which may have led to the loss of many features. Conserved features that were specific to Xtg were identified, and further investigation will help to determine genes that are essential to pathogenicity and host adaptation of Xtg.

plant biology↗

bioRxiv2022

Phenotypic variation and quantitative trait loci for resistance to southern anthracnose and clover rot in red clover

Red clover (Trifolium pratense L.) is an important forage legume of temperate regions, particularly valued for its high yield potential and its high forage quality. Despite substantial breeding progress during the last decades, continuous improvement of cultivars is crucial to ensure yield stability in view of newly emerging diseases or changing climatic conditions. The high amount of genetic diversity present in red clover ecotypes, landraces and cultivars provides an invaluable, but often unexploited resource for the improvement of key traits such as yield, quality, and resistance to biotic and abiotic stresses. A collection of 397 red clover accessions was genotyped using a pooled genotyping-by-sequencing approach with 200 plants per accession. Resistance to the two most pertinent diseases in red clover production, southern anthracnose caused by Colletotrichum trifolii, and clover rot caused by Sclerotinia trifoliorum, was assessed using spray inoculation. The mean survival rate for southern anthracnose was 22.9% and the mean resistance index for clover rot was 34.0%. Genome-wide association analysis revealed several loci significantly associated with resistance to southern anthracnose and clover rot. Most of these loci are in coding regions. One quantitative trait locus (QTL) on chromosome 1 explained 16.8% of the variation in resistance to southern anthracnose. For clover rot resistance we found eight QTL, explaining together 80.2% of the total phenotypic variation. The SNPs associated with these QTL provide, once validated, a promising resource for marker-assisted selection in existing breeding programs, facilitating the development of novel cultivars with increased resistance against two devastating fungal diseases of red clover. Key messageHigh variability for and candidate loci associated with resistance to southern anthracnose and clover rot in a worldwide collection of red clover provide a first basis for genomics-assisted breeding.

plant biology↗

bioRxiv2021

A multispecies amplicon sequencing approach for genetic diversity assessment in grassland plant species

Grasslands are widespread and economically relevant ecosystems at the basis of sustainable roughage production. Plant genetic diversity (PGD; i.e., within-species diversity) is related to many beneficial effects to the ecosystem functioning of grasslands. The monitoring of PGD in temperate grasslands is complicated by the multiplicity of species present and by a shortage of methods for large-scale assessment. However, the continuous advancement of high-throughput DNA sequencing approaches have improved the prospects of broad, multispecies PGD monitoring. Among them, amplicon sequencing stands out as a robust and cost-effective method. Here we report a set of twelve multispecies primer pairs that can be used for high-throughput PGD assessment in multiple grassland plant species. The loci targeted by the amplicons were selected and tested in two phases: a "discovery phase" based on a sequence capture assay (611 target nuclear loci assessed in 16 grassland plant species), which resulted in the selection of eleven loci; and a "validation phase", in which the selected loci were targeted and sequenced using twelve multispecies primers in test populations of Dactylis glomerata L., Lolium perenne L., Festuca pratensis Huds., Trifolium pratense L. and T. repens L. The resulting multispecies amplicons had overall nucleotide diversities per species ranging from 5.19 x 10-3 to 1.29 x 10-2, which is in the range of flowering-related genes but slightly lower than pathogen resistance genes. We conclude that the methodology, the DNA sequence resources, and the amplicon-specific primer pairs reported in this study provide the basis for large-scale, multispecies PGD monitoring in grassland plants.

ecology↗

bioRxiv2021

Evidence for high intergenic sequence variation in heterozygous Italian ryegrass (Lolium multiflorum Lam.) genome revealed by a high-quality draft diploid genome assembly

BackgroundOver the last decade, progress in DNA sequencing technologies and assembly methods allowed plant scientists to move beyond the use of model organisms and work directly on the genomes of the major crops. Forage grass research can also benefit from this revolution, enabling progress in population genetic studies, functional biology, and genomics-assisted breeding. Due to its large genome size and high repeat content, so far only incomplete and fragmented assemblies are available for the grasses of the Lolium and Festuca species complex. FindingsHere, we report a highly contiguous draft assembly of Italian ryegrass (L. multiflorum Lam.), spanning 4.5 Gb and with a N50 of 3 Mb, containing ~70,000 gene models. Thanks to its relatedness to barley, 78% of the assembly was anchored on seven pseudomolecules. The high heterozygosity of the plant allowed obtaining a diploid assembly - i.e. across 95% of the assembly, both alleles were assembled on separate sequences. This feature allowed unraveling a very high amount of intergenic sequence variation between allelic sequences. ConclusionsWe present a nearly complete genome assembly of a genotype used in contemporary Swiss forage grass breeding programs. This work shows how genomic research has improved, allowing to decode the genetic code of large, complex, and heterozygous plants. It also allows the functional characterization of the ryegrass gene repertoire and the large-scale development of molecular markers. Furthermore, it paves the road for a reference-based characterization and exploitation of the genetic variation within the Lolium and Festuca species complex.

genomics↗