TY  - JOUR
AB  - Plant defensins (PDFs) are cysteine-rich antimicrobial peptides (AMPs) that are important components of plant immunity. They occur constitutively in various plant tissues but are also upregulated upon stress. Therefore, these molecules are of great interest as markers for the diagnosis of early forest stress response in plants at the molecular level. PDFs are small peptides (~5 kDa) with a compact tertiary structure, requiring specific protocols and dedicated antibodies for detection by quantitative ELISA. We developed monoclonal recombinant antibodies using phage display in solution against the correctly folded antigen defensin FsPDF2 from beech (Fagus sylvatica) and analysed the antibody-antigen interaction in silico with AlphaFold 3. In a proof-of-principle study, we investigated the FsPDF2 stress response to abiotic (drought) and biotic (gall midge) stresses. Notably, we established an assay for defensin quantification in crude plant extract, detecting for the first time natively folded proteins in a specific sandwich ELISA. Our antibody generation strategy can be transferred by practitioners to other small antimicrobial peptides (AMP), paving the way to study this group of proteins and their corresponding stress response comprehensively.
AU  - Heine, P.A.*
AU  - Nosenko, T.
AU  - Kistner, S.*
AU  - Oliphant, K.D.*
AU  - Hanke-Uhe, M.*
AU  - Shahid, A.*
AU  - Hu, B.*
AU  - Kucklick, M.*
AU  - Lehmler, N.*
AU  - Becker, M.*
AU  - Goerke, N.*
AU  - Korn, J.M.*
AU  - Linke, T.*
AU  - Meier, D.H.*
AU  - Perl, A.*
AU  - Polten, S.*
AU  - Priess, V.*
AU  - Schäckermann, D.*
AU  - Schubert, M.*
AU  - Schumacher, J.*
AU  - Winkler, J.B.
AU  - Engelmann, S.*
AU  - Rennenberg, H.*
AU  - Schnitzler, J.-P.
AU  - Dübel, S.*
AU  - Hust, M.*
AU  - Hänsch, R.*
AU  - Kaufholdt, D.*
C1  - 75994
C2  - 58334
TI  - Phage display derived antibodies against antimicrobial peptide FsPDF2 reveal stress response in European beech.
JO  - Plant Biotechnol. J.
PY  - 2025
SN  - 1467-7644
ER  - 

TY  - JOUR
AB  - Nuclear magnetic resonance (NMR) spectroscopy can be used to determine the structure, dynamics and interactions of proteins. However, protein NMR requires stable isotope labeling for signal detection. The cells used for the production of recombinant proteins must therefore be grown in medium containing isotopically labeled substrates. Stable isotope labeling is well established in Escherichia coli, but bacteria are only suitable for the production of simple proteins without post-translational modifications. More complex proteins require eukaryotic production hosts, but their growth can be impaired by labeled media, thus reducing product yields and increasing costs. To address this limitation, we used media supplemented with isotope-labeled substrates to cultivate the tobacco-derived cell line BY-2, which was then cast into plant cell packs (PCPs) for the transient expression of a labeled version of the model protein GB1. Mass spectrometry confirmed the feasibility of isotope labeling with 15 N and 2 H using this approach. The resulting NMR spectrum featured a signal dispersion comparable to recombinant GB1 produced in E. coli. PCPs therefore offer a rapid and cost-efficient alternative for the production of isotope-labeled proteins for NMR analysis, especially suitable for complex proteins that cannot be produced in microbial systems.
AU  - Opdensteinen, P.*
AU  - Sperl, L.E.
AU  - Mohamadi, M.
AU  - Kündgen-Redding, N.*
AU  - Hagn, F.
AU  - Buyel, J.F.*
C1  - 65427
C2  - 52292
SP  - 1928-1939
TI  - The transient expression of recombinant proteins in plant cell packs facilitates stable isotope labeling for NMR spectroscopy.
JO  - Plant Biotechnol. J.
VL  - 20
IS  - 10
PY  - 2022
SN  - 1467-7644
ER  - 

TY  - JOUR
AB  - The capacity of the bread wheat (Triticum aestivum) genome to tolerate introgression from related genomes can be exploited for wheat improvement. A resistance to powdery mildew expressed by a derivative of the cross-bread wheat cv. Tähti × T. militinae (Tm) is known to be due to the incorporation of a Tm segment into the long arm of chromosome 4A. Here, a newly developed in silico method termed rearrangement identification and characterization (RICh) has been applied to characterize the introgression. A virtual gene order, assembled using the GenomeZipper approach, was obtained for the native copy of chromosome 4A; it incorporated 570 4A DArTseq markers to produce a zipper comprising 2132 loci. A comparison between the native and introgressed forms of the 4AL chromosome arm showed that the introgressed region is located at the distal part of the arm. The Tm segment, derived from chromosome 7G, harbours 131 homoeologs of the 357 genes present on the corresponding region of Chinese Spring 4AL. The estimated number of Tm genes transferred along with the disease resistance gene was 169. Characterizing the introgression's position, gene content and internal gene order should not only facilitate gene isolation, but may also be informative with respect to chromatin structure and behaviour studies.
AU  - Abrouk, M.*
AU  - Balcárková, B.*
AU  - Šimková, H.*
AU  - Komínkova, E.*
AU  - Martis, M.M.
AU  - Jakobson, I.*
AU  - Timofejeva, L.*
AU  - Rey, E.*
AU  - Vrana, J.*
AU  - Kilian, A.*
AU  - Järve, K.*
AU  - Dolezel, J.*
AU  - Valárik, M.*
C1  - 49553
C2  - 40704
CY  - Hoboken
SP  - 249-256
TI  - The in silico identification and characterization of a bread wheat/Triticum militinae introgression line.
JO  - Plant Biotechnol. J.
VL  - 15
IS  - 2
PB  - Wiley-blackwell
PY  - 2017
SN  - 1467-7644
ER  - 

TY  - JOUR
AB  - Fusarium head blight is a devastating disease of small grain cereals such as bread wheat (Triticum aestivum). The pathogen switches from a biotrophic to a nectrotrophic lifestyle in course of disease development forcing its host to adapt its defence strategies. Using a genetical genomics approach we illustrate genome-wide reconfigurations of genetic control over transcript abundances between two decisive time points after inoculation with the causative pathogen Fusarium graminearum. Whole transcriptome measurements have been recorded for 163 lines of a wheat doubled haploid population segregating for several resistance genes yielding 15 552 at 30 hours and 15 888 eQTL at 50 hours after inoculation. The genetic map saturated with transcript abundance-derived markers identified of a novel QTL on chromosome 6A, besides the previously reported QTL Fhb1 and Qfhs.ifa-5A. We find a highly different distribution of eQTL between time points with about 40% of eQTL being unique for the respective assessed time points. But also for more than 20% of genes governed by eQTL at either time point genetic control changes in time. These changes are reflected in the dynamic compositions of three major regulatory hotspots on chromosomes 2B, 4A and 5A. In particular control of defence-related biological mechanisms concentrated in the hotspot at 4A shift to hotspot 2B as the disease progresses. Hotspots do not colocalize with phenotypic QTL and within their intervals no higher than expected number of eQTL was detected. Thus, resistance conferred by either QTL is mediated by few or single genes.
AU  - Samad-Zamini, M.*
AU  - Schweiger, W.*
AU  - Nussbaumer, T.
AU  - Mayer, K.F.X.
AU  - Buerstmayr, H.*
C1  - 50784
C2  - 42871
CY  - Hoboken
SP  - 1453–1464
TI  - Time-course expression QTL atlas of the global transcriptional response of wheat to Fusarium graminearum.
JO  - Plant Biotechnol. J.
VL  - 15
IS  - 11
PB  - Wiley
PY  - 2017
SN  - 1467-7644
ER  - 

TY  - JOUR
AB  - Hierarchical shotgun sequencing remains the method of choice for assembling high-quality reference sequences of complex plant genomes. The efficient exploitation of current high-throughput technologies and powerful computational facilities for large-insert clone sequencing necessitates the sequencing and assembly of a large number of clones in parallel. We developed a multiplexed pipeline for shotgun sequencing and assembling individual bacterial artificial chromosomes (BACs) using the Illumina sequencing platform. We illustrate our approach by sequencing 668 barley BACs (Hordeum vulgare L.) in a single Illumina HiSeq 2000 lane. Using a newly designed parallelized computational pipeline, we obtained sequence assemblies of individual BACs that consist, on average, of eight sequence scaffolds and represent >98% of the genomic inserts. Our BAC assemblies are clearly superior to a whole-genome shotgun assembly regarding contiguity, completeness and the representation of the gene space. Our methods may be employed to rapidly obtain high-quality assemblies of a large number of clones to assemble map-based reference sequences of plant and animal species with complex genomes by sequencing along a minimum tiling path.
AU  - Beier, S.*
AU  - Himmelbach, A.*
AU  - Schmutzer, T.*
AU  - Felder, M.*
AU  - Taudien, S.*
AU  - Mayer, K.F.X.
AU  - Platzer, M.*
AU  - Stein, N.*
AU  - Scholz, U.*
AU  - Mascher, M.*
C1  - 47741
C2  - 39485
CY  - Hoboken
SP  - 1511-1522
TI  - Multiplex sequencing of bacterial artificial chromosomes for assembling complex plant genomes.
JO  - Plant Biotechnol. J.
VL  - 14
IS  - 7
PB  - Wiley-blackwell
PY  - 2016
SN  - 1467-7644
ER  - 

TY  - JOUR
AB  - Two Gram-negative, plant growth-promoting rhizobacteria (PGPRs), denominated as M12 and M14, were classified by 16S rDNA sequencing as Burkholderia graminis species. Both strains were shown to produce a variety of N-acyl-homoserine lactone (AHL) quorum sensing (QS) signalling molecules. The involvement of these molecules in plant growth promotion and the induction of protection against salt stress was examined. AHL production was evaluated in vitro by thin-layer chromatography using AHL biosensors, and the identity of the AHLs produced was determined by liquid chromatography-tandem mass spectrometry. The in situ production of AHLs by M12 and M14 in the rhizosphere of Arabidopsis thaliana plants was detected by co-inoculation with green fluorescent protein-based biosensor strains and confocal laser scanning microscopy. To determine whether plant growth promotion and protection against salt stress were mediated by QS, these PGPRs were assayed on wild-type tomato plants, as well as their corresponding transgenics expressing YenI (short-chain AHL producers) and LasI (long-chain AHL producers). In wild-type tomato plants, only M12 promoted plant growth, and this effect disappeared in both transgenic lines. In contrast, M14 did not promote growth in wild-type tomatoes, but did so in the LasI transgenic line. Resistance to salt stress was induced by M14 in wild-type tomato, but this effect disappeared in both transgenic lines. The strain M12, however, did not induce salt resistance in wild-type tomato, but did so in LasI tomato plants. These results reveal that AHL QS signalling molecules mediate the ability of both PGPR strains M12 and M14 to promote plant growth and to induce protection against salt stress.
AU  - Barriuso, J.*
AU  - Ramos Solano, B.*
AU  - Fray, R.G.*
AU  - Cámara, M.*
AU  - Hartmann, A.
AU  - Gutiérrez Manero, F.J.*
C1  - 1267
C2  - 25813
SP  - 442-452
TI  - Transgenic tomato plants alter quorum sensing in plant growth-promoting rhizobacteria.
JO  - Plant Biotechnol. J.
VL  - 6
IS  - 5
PB  - Blackwell
PY  - 2008
SN  - 1467-7644
ER  -