Biology subjects

Dunemann, F.

Publications and source records attributed to Dunemann, F..

5 recordsLinked to original sources

bioRxiv2025

Endogenous falcarinol-type polyacetylenes in carrots and their putative influence on post-harvest fungal pathogens Mycocentrospora acerina and Botrytis cinerea

Falcarinol-type polyacetylenes are a main group of natural bitter compounds synthesized in carrot taproots and putatively contribute to defence mechanisms against soil-borne fungal pathogens. In this study, we analysed the relationships between the constitutive levels of these secondary metabolites and the extent of root infection with the two main carrot storage fungal pathogens Mycocentrospora acerina (liquorice rot) and Botrytis cinerea (grey mold). Taproots of eight differently colored carrot cultivars exhibiting different levels of the three main falcarinol-type polyacetylenes were inoculated with the two fungi and evaluated for diseased area with a digital image analysis system after 6-weeks incubation in a cold storage facility. After inoculation on cortex tissue and the periderm of taproots, a progressed carrot breeding line and the cultivar Bolero demonstrated the highest tolerance to both M. acerina and B. cinerea, while Blanche and Senta were the most susceptible cultivars. Concentrations of falcarinol-type polyacetylenes were negatively correlated, though not significantly, with M. acerina and B. cinerea disease severity. Nevertheless, our study has provided further insights into the naturally occurring variability for the accumulation of falcarinol-type polyacetylenes in cultivated carrots and their putative contributions to resistance against post-harvest fungal pathogens M. acerina and B. cinerea. To our knowledge, this is the first study about the pathogenicity of B. cinerea after inoculation of different carrot cultivars with known root PA levels.

plant biology↗

bioRxiv2022

Plasticity in centromere organization: Holocentromeres can consist of merely a few megabase-sized satellite arrays

The centromere is the chromosome region where the microtubules attach during cell division. In contrast to monocentric chromosomes with one centromere location, holocentric species usually distribute hundreds of centromere units along the entire chromatid. We assembled the chromosome-scale reference genome and analyzed the holocentromere and (epi)genome organization of the lilioid Chionographis japonica. Remarkably, each of its holocentric chromatids consists of only 7 to 11 evenly-spaced megabase-sized centromere-specific histone H3-positive units. These units contain satellite arrays of 23 and 28 bp-long monomers capable of forming palindromic structures. Like monocentric species, C. japonica forms distinctly clustered centromeres in chromocenters at interphase. Additionally, the large-scale eu- and heterochromatin arrangement differs between C. japonica and other known holocentric species. Using polymer simulations, we modeled the formation of prometaphase line-like holocentromeres from interphase centromere clusters. Our findings broaden the knowledge about the diversity of centromere organization, showing that holocentricity is not restricted to species with numerous and small centromere units.

plant biology↗

bioRxiv2022

Genome-wide identification of putative disease resistance genes (R genes) in carrot (Daucus carota subsp. sativus) by homology-based gene prediction

The cultivated carrot (Daucus carota ssp. sativus) is one of the most important root vegetable crops grown worldwide. Carrots are highly susceptible to several pests and diseases, and disease resistance is currently among the main breeding aims. The inheritance of resistance has been reported for a few carrot foliar diseases and root-knot nematodes, but no functionally characterized resistance gene (R gene) has yet been linked as a candidate gene to any resistance locus in carrot. Knowledge about the inventory of NLR genes (nucleotide-binding leucine-rich repeat receptors) and other R genes encoding transmembrane proteins such as receptor-like proteins (RLPs) and kinases (KIN) would be necessary to associate major QTLs (quantitative trait loci) identified by bi-parental QTL analyses or GWAS (genome-wide association analysis) with functional candidate R genes. In this study, we describe a combination of a genome-wide inventory of putative full-length carrot R genes based on a homology-based gene prediction approach called GeMoMa and subsequent classification by usage of the recent version of PRGdb 4.0 database (Calle-Garcia et al. 2022). A total of 320 putative carrot R genes were identified and bioinformatically characterized, including 72 newly identified gene models, that have not yet been annotated in the currently available carrot whole genome sequence. Based on the DRAGO 3 pipeline, totally 137 putative NLR genes were found, whereas 162 putative functional RLP and KIN genes were identified in the carrot genome. About one third of the R genes was found to be organized in clusters consisting only of NLR, RLP or KIN genes. To determine the evolutionary relationships of carrot R gene predictions, we generated a phylogenetic tree based on the alignment of all 320 R proteins. Three large clades (NLR, RLP and KIN) and a small clade (RLK) were identified, which reflect well the classification obtained after DRAGO 3 analysis. The presented carrot R gene inventory might be useful for resistance gene isolation, the development of (functional) molecular markers and resistance breeding in carrot.

plant biology↗

bioRxiv2022

Insights from CRISPR/Cas9-mediated gene editing of centromeric histone H3 (CENH3) in carrot (Daucus carota subsp. sativus)

The generation of haploids is one of the most powerful means to accelerate the plant breeding process. In most crop species, an efficient haploid technology is not yet available or only applicable to a limited set of genotypes. Recent results published for Arabidopsis thaliana and major cereal crops like maize and wheat about successful haploid induction by CRISPR/Cas9-mediated editing of the centromeric histone H3 gene (CENH3) suggest that this novel method for the production of haploid plants might also be applicable to vegetable species like carrot. Here, we report and summarize the different experimental and genetic approaches that have been focused in the past few years on CRISPR/Cas9-based editing of the carrot CENH3 gene. We also describe the discovery of a second CENH3 locus in the carrot genome, which complicates the attempts to generate and to analyse putative haploid inducer genotypes. We show that three different CRISPR/Cas9 target constructs, used alone or in combinations, could successfully target carrot CENH3. Promising mutants such as in-frame indel or in-frame deletion mutants have been found, but their successful usage as putative haploid inducer is uncertain yet. Next generation sequencing of amplicons spanning CRISPR target sites and transcript-based amplicon sequencing seemed to be appropriate methods to select promising mutants, to estimate mutation frequencies, and to allow a first prediction which gene was concerned. Another aim of this study was the simultaneous knockout and complementation of the endogenous carrot CENH3 gene by an alien CENH3 gene. Co-transformation of a CRISPR/Cas9-based carrot CENH3 knockout construct together with a CENH3 gene cloned from ginseng (Panax ginseng) was performed by using Rhizobium rhizogenes. It was shown, that ginseng CENH3 protein is accumulated inside the kinetochore region of carrot chromosomes, indicating that PgCENH3 might be a suited candidate for this approach. However, presently it is unclear, if this gene is fully functioning during the meiotic cell divisions and able to complement lethal gametes. Challenges and future prospects to develop a CENH3-based HI system for carrot are discussed.

plant biology↗

bioRxiv2020

Application of aptamers improves CRISPR-based live imaging of plant telomeres

Development of live imaging techniques for providing information how chromatin is organized in living cells is pivotal to decipher the regulation of biological processes. Here, we demonstrate the improvement of a live imaging technique based on CRISPR/Cas9. In this approach, the sgRNA scaffold is fused to RNA aptamers including MS2 and PP7. When the dead Cas9 (dCas9) is co-expressed with chimeric sgRNA, the aptamer-binding proteins fused to fluorescent protein (MCP-FP and PCP-FP) are recruited to the targeted sequence. Compared to previous work with dCas9:GFP, we show that the quality of telomere labelling was improved in transiently transformed Nicotiana benthamiana using aptamer-based CRISPR-imaging constructs. Labelling is influenced by the copy number of aptamers and less by the promoter types. The same constructs were not applicable for labelling of repeats in stably transformed plants and roots. The constant interaction of the RNP complex with its target DNA might interfere with cellular processes. HighlightAptamer-based CRISPR imaging: an opportunity for improving live-cell imaging in plants

cell biology↗