This study identified the phenotypic and genotypic characteristics of the bacteria that nodulate wild Lathyrus and Vicia species natural distribution in the Gaziantep province of Turkey. Principle component analysis of phenotypic features revealed that rhizobial isolates were highly resistant to stress factors such as high salt, pH and temperature. They were found to be highly sensitive to the concentrations (mg/mL) of the antibiotics neomycin 10, kanamycin, and tetracycline 5, as well as the heavy metals Ni 10, and Cu 10, and 5. As a result of REP-PCR analysis, it was determined that the rhizobial isolates were quite diverse, and 5 main groups and many subgroups being found. All of the isolates nodulating wild Vicia species were found to be related to Rhizobium sp., and these isolates were found to be in Clades II, III, IV, and V of the phylogenetic tree based on 16S rRNA. The isolates that nodulated wild Lathyrus species were in Clades I, II, IV, V, VI, VII, and VIII, and they were closely related to Rhizobium leguminasorum, Rhizobium sp., Phyllobacterium sp., Serratia sp., and Pseudomonas sp. According to the genetic analyses, the isolates could not be classified at the species level, the similarity ratio was low, they formed a distinct group that was supported by strong bootstrap values in the phylogenetic tree, and the differences discovered in the network analysis revealed the diversity among the isolates and gave important findings that these isolates may be new species.
- MeSH
- antibakteriální látky farmakologie MeSH
- Bacteria genetika klasifikace izolace a purifikace účinky léků MeSH
- DNA bakterií genetika MeSH
- fenotyp MeSH
- fylogeneze * MeSH
- genetická variace MeSH
- genotyp * MeSH
- kořenové hlízky rostlin * mikrobiologie MeSH
- Lathyrus * mikrobiologie MeSH
- Rhizobium genetika klasifikace izolace a purifikace MeSH
- RNA ribozomální 16S * genetika MeSH
- těžké kovy MeSH
- vikev * mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Turecko MeSH
The Venus flytrap (Dionaea muscipula J. Ellis) is a carnivorous plant able to synthesize large amounts of phenolic compounds, such as phenylpropanoids, flavonoids, phenolic acids, and 1,4-naphtoquinones. In this study, the first genetic transformation of D. muscipula tissues is presented. Two wild-type Rhizobium rhizogenes strains (LBA 9402 and ATCC 15834) were suitable vector organisms in the transformation process. Transformation led to the formation of teratoma (transformed shoot) cultures with the bacterial rolB gene incorporated into the plant genome in a single copy. Using high-pressure liquid chromatography, we demonstrated that transgenic plants were characterized by an increased quantity of phenolic compounds, including 1,4-naphtoquinone derivative, plumbagin (up to 106.63 mg × g-1 DW), and phenolic acids (including salicylic, caffeic, and ellagic acid), in comparison to non-transformed plants. Moreover, Rhizobium-mediated transformation highly increased the bactericidal properties of teratoma-derived extracts. The antibacterial properties of transformed plants were increased up to 33% against Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli and up to 7% against Pseudomonas aeruginosa. For the first time, we prove the possibility of D. muscipula transformation. Moreover, we propose that transformation may be a valuable tool for enhancing secondary metabolite production in D. muscipula tissue and to increase bactericidal properties against human antibiotic-resistant bacteria. KEY POINTS: • Rhizobium-mediated transformation created Dionaea muscipula teratomas. • Transformed plants had highly increased synthesis of phenolic compounds. • The MBC value was connected with plumbagin and phenolic acid concentrations.
- MeSH
- Agrobacterium genetika MeSH
- antibakteriální látky farmakologie MeSH
- Droseraceae * MeSH
- fenoly MeSH
- lidé MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Nanotechnology, new fascinating field of science, is bringing many application's options. However, it is necessary to understand their potential environmental risk and toxicity. Zinc selenide quantum dots (ZnSe QDs) are getting valuable due to wide industrial usage, mainly as cadmium free diodes or stabilizing ligand. Thanks to unique properties, they could also open the possibilities of application in the agriculture. Their effects on living organisms, including plants, are still unknown. Therefore, the attention of this work was given to antioxidant response of Arabidopsis thaliana to 100 and 250 μM ZnSe QDs foliar feeding. ZnSe QDs treatment had no statistically significant differences in morphology but led to increased antioxidant response in the leaves at the level of gene expression and production secondary antioxidant metabolites. Concurrently, analysis of growth properties of Agrobacterium tumefaciens was done. 250 μM ZnSe solution inhibited the Agrobacterium tumefaciens viability by 60%. This is the first mention about effect ZnSe QDs on the plants. Although QDs induced oxidative stress, the apply treatment dose of ZnSe QDs did not have significant toxic effect on the plants and even no morphological changes were observed. However, the same amount of ZnSe QD induced an inhibitory effect on Agrobacterium tumefaciens.
- MeSH
- Agrobacterium tumefaciens účinky léků růst a vývoj MeSH
- antioxidancia metabolismus MeSH
- Arabidopsis genetika růst a vývoj metabolismus MeSH
- exprese genu * MeSH
- kvantové tečky toxicita MeSH
- metabolomika * MeSH
- sloučeniny selenu aplikace a dávkování toxicita MeSH
- sloučeniny zinku aplikace a dávkování toxicita MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Plant-rhizobia symbiosis can activate key genes involved in regulating nodulation associated with biological nitrogen fixation (BNF). Although the general molecular basis of the BNF process is frequently studied, little is known about its intraspecific variability and the characteristics of its allelic variants. This study's main goals were to describe phenotypic and genotypic variation in the context of nitrogen fixation in red clover (Trifolium pretense L.) and identify variants in BNF candidate genes associated with BNF efficiency. Acetylene reduction assay validation was the criterion for selecting individual plants with particular BNF rates. Sequences in 86 key candidate genes were obtained by hybridization-based sequence capture target enrichment of plants with alternative phenotypes for nitrogen fixation. Two genes associated with BNF were identified: ethylene response factor required for nodule differentiation (EFD) and molybdate transporter 1 (MOT1). In addition, whole-genome population genotyping by double-digest restriction-site-associated sequencing (ddRADseq) was performed, and BNF was evaluated by the natural 15N abundance method. Polymorphisms associated with BNF and reflecting phenotype variability were identified. The genetic structure of plant accessions was not linked to BNF rate of measured plants. Knowledge of the genetic variation within BNF candidate genes and the characteristics of genetic variants will be beneficial in molecular diagnostics and breeding of red clover.
- MeSH
- alely MeSH
- fenotyp MeSH
- fixace dusíku genetika MeSH
- genotyp MeSH
- interakce mikroorganismu a hostitele MeSH
- kořeny rostlin genetika mikrobiologie MeSH
- polymorfismus genetický * MeSH
- Rhizobium fyziologie MeSH
- rostlinné geny genetika MeSH
- sekvenční analýza DNA metody MeSH
- symbióza genetika MeSH
- Trifolium genetika mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
Phytohormones, similar to soil enzymes, are synthesized and secreted into the soil environment by fungi and microorganisms. Phytohormones are involved in regulating microbial community activity in the rhizosphere. This paper examines how auxins, cytokinins, ethephon and chlorocholine chloride affect the activity of native soil proteases in the organo-mineral horizon of an alpine meadow. In the meadow habitat, native soil proteases were inhibited by auxins whereas the effect of cytokinins on these enzymes was not statistically significant. A similar inhibitory effect on the activity of proteases was shown for ethephon and chlorocholine chloride, both of which also inhibited the activity of native soil proteases in the alpine meadow soil. Overall, the inhibitory effect of phytohormones on the activity of native protease activity may affect plant nutrition by retarding the nitrogen cycle in the soil. This work contributes to our understanding of the influence of substances produced by the rhizosphere that can actively participate in the activity of soil microorganisms and consequently influence the soil nitrogen cycle.
- MeSH
- chlormekvát farmakologie MeSH
- cytokininy farmakologie MeSH
- dusík chemie MeSH
- kyseliny indoloctové farmakologie MeSH
- organofosforové sloučeniny farmakologie MeSH
- pastviny MeSH
- proteasy metabolismus MeSH
- půdní mikrobiologie MeSH
- regulátory růstu rostlin farmakologie MeSH
- Rhizobiaceae enzymologie MeSH
- rostlinné proteiny metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Many Gram-negative bacteria use N-acyl-homoserine lactones (N-acyl-HSL) as quorum sensing signal molecules. N-acyl-HSL are the subject of investigation by many research groups. Simple methods have been designed to detect N-acyl-HSL using bacterial strains that doesn't produce its own signal molecules, but the reporter gene is expressed when exogenous N-acyl-HSL are present. This article reviews and discusses the most common and most currently obtainable systems using bacteria such as Agrobacterium tumefaciens, Chromobacterium violaceum, Escherichia coli or Vibrio fischeri. These biological sensing elements can be used to detect and study a wide range of N-acyl-HSL and related molecules interfering with quorum sensing systems.
Organophosphorus compounds have been widely employed to the development of warfare nerve agents and pesticides, resulting in a huge number of people intoxicated annually, being a serious problem of public health. Efforts worldwide have been done in order to design new technologies that are capable of combating or even reversing the poisoning caused by these OP nerve agents. In this line, the bioremediation arises as a promising and efficient alternative for this purpose. As an example of degrading enzymes, there is the organophosphate-degrading (OpdA) enzyme from Agrobacterium radiobacter, which has been quite investigated experimentally due to its high performance in the degradation of neurotoxic nerve agents. This work aims to look into the structural and electronic details that govern the interaction modes of these compounds in the OpdA active site, with the posterior hydrolysis reaction prediction. Our findings have brought about data about the OpdA performance towards different nerve agents, and among them, we may realize that the degradation efficiency strongly depends on the nerve agent structure and its stereochemistry, being in this case the compound Tabun the one more effectively hydrolyzed. By means of the chemical bonds (AIM) and orbitals (FERMO) analysis, it is suggested that the initial reactivity of the OP nerve agents in the OpdA active site does not necessarily dictate the reactivity and interaction modes over the reaction coordinate.
- MeSH
- Agrobacterium tumefaciens enzymologie MeSH
- bakteriální proteiny chemie metabolismus MeSH
- biodegradace * MeSH
- biokatalýza MeSH
- fosfatasy chemie metabolismus MeSH
- katalytická doména MeSH
- kvantová teorie MeSH
- lidé MeSH
- nervová bojová látka chemie metabolismus MeSH
- sarin chemie metabolismus MeSH
- simulace molekulového dockingu MeSH
- vazebná místa MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Silene latifolia serves as a model species to study dioecy, the evolution of sex chromosomes, dosage compensation and sex-determination systems in plants. Currently, no protocol for genetic transformation is available for this species, mainly because S. latifolia is considered recalcitrant to in vitro regeneration and infection with Agrobacterium tumefaciens. Using cytokinins and their synthetic derivatives, we markedly improved the efficiency of regeneration. Several agrobacterial strains were tested for their ability to deliver DNA into S. latifolia tissues leading to transient and stable expression of the GUS reporter. The use of Agrobacterium rhizogenes strains resulted in the highest transformation efficiency (up to 4.7% of stable transformants) in hairy root cultures. Phenotypic and genotypic analyses of the T1 generation suggested that the majority of transformation events contain a small number of independent T-DNA insertions and the transgenes are transmitted to the progeny in a Mendelian pattern of inheritance. In short, we report an efficient and reproducible protocol for leaf disc transformation and subsequent plant regeneration in S. latifolia, based on the unique combination of infection with A. rhizogenes and plant regeneration from hairy root cultures using synthetic cytokinins. A protocol for the transient transformation of S.latifolia protoplasts was also developed and applied to demonstrate the possibility of targeted mutagenesis of the sex linked gene SlAP3 by TALENs and CRISPR/Cas9.
- MeSH
- Agrobacterium genetika MeSH
- chromozomy rostlin genetika MeSH
- CRISPR-Cas systémy MeSH
- DNA bakterií genetika MeSH
- exprese genu MeSH
- genetické techniky MeSH
- geneticky modifikované rostliny MeSH
- modely genetické MeSH
- molekulární evoluce MeSH
- regenerace genetika MeSH
- reportérové geny MeSH
- Silene genetika mikrobiologie fyziologie MeSH
- TALENs MeSH
- transformace genetická * MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Plant transformation via Agrobacterium tumefaciens is characterized by integration of commonly low number of T-DNAs at random positions in the genome. When integrated into an active gene region, promoterless reporter genes placed near the T-DNA border sequence are frequently transcribed and even translated to reporter proteins, which is the principle of promoter- and gene-trap lines. RESULTS: Here we show that even internal promotorless regions of T-DNAs are often transcribed. Such spontaneous transcription was observed in the majority of independently transformed tobacco BY-2 lines (over 65%) and it could effectively induce silencing if an inverted repeat was present within the T-DNA. We documented that the transcription often occurred in both directions. It was not directly connected with any regulatory elements present within the T-DNAs and at least some of the transcripts were initiated outside of the T-DNA. The likeliness of this read-through transcription seemed to increase in lines with higher T-DNA copy number. Splicing and presence of a polyA tail in the transcripts indicated involvement of Pol II, but surprisingly, the transcription was able to run across two transcription terminators present within the T-DNA. Such pervasive transcription was observed with three different T-DNAs in BY-2 cells and with lower frequency was also detected in Arabidopsis thaliana. CONCLUSIONS: Our results demonstrate unexpected pervasive read-through transcription of T-DNAs. We hypothesize that it was connected with a specific chromatin state of newly integrated DNA, possibly affected by the adjacent genomic region. Although this phenomenon can be easily overlooked, it can have significant consequences when working with highly sensitive systems like RNAi induction using an inverted repeat construct, so it should be generally considered when interpreting results obtained with the transgenic technology.
- MeSH
- Agrobacterium tumefaciens genetika MeSH
- Arabidopsis genetika MeSH
- buněčné linie MeSH
- DNA bakterií genetika MeSH
- genetická transkripce * MeSH
- geneticky modifikované rostliny MeSH
- messenger RNA genetika MeSH
- obrácené repetice genetika MeSH
- promotorové oblasti (genetika) genetika MeSH
- reportérové geny MeSH
- tabák genetika MeSH
- Publikační typ
- časopisecké články MeSH
Old Yellow Enzymes (OYEs) are NAD(P)H dehydrogenases of not fully resolved physiological roles that are widespread among bacteria, plants, and fungi and have a great potential for biotechnological applications. We determined the apo form crystal structure of a member of the OYE class, glycerol trinitrate reductase XdpB, from Agrobacterium bohemicum R89-1 at 2.1 Å resolution. In agreement with the structures of the related bacterial OYEs, the structure revealed the TIM barrel fold with an N-terminal β-hairpin lid, but surprisingly, the structure did not contain its cofactor FMN. Its putative binding site was occupied by a pentapeptide TTSDN from the C-terminus of a symmetry related molecule. Biochemical experiments confirmed a specific concentration-dependent oligomerization and a low FMN content. The blocking of the FMN binding site can exist in vivo and regulates enzyme activity. Our bioinformatic analysis indicated that a similar self-inhibition could be expected in more OYEs which we designated as subgroup OYE C1. This subgroup is widespread among G-bacteria and can be recognized by the conserved sequence GxxDYP in proximity of the C termini. In proteobacteria, the C1 subgroup OYEs are typically coded in one operon with short-chain dehydrogenase. This operon is controlled by the tetR-like transcriptional regulator. OYEs coded in these operons are unlikely to be involved in the oxidative stress response as the other known members of the OYE family because no upregulation of XdpB was observed after exposing A. bohemicum R89-1 to oxidative stress.
- MeSH
- Agrobacterium enzymologie genetika MeSH
- bakteriální geny MeSH
- bakteriální proteiny chemie genetika metabolismus MeSH
- flavinmononukleotid metabolismus MeSH
- katalytická doména MeSH
- kinetika MeSH
- krystalografie rentgenová MeSH
- kvarterní struktura proteinů MeSH
- molekulární modely MeSH
- NADPH-dehydrogenasa chemie genetika metabolismus MeSH
- operon MeSH
- oxidační stres MeSH
- oxidoreduktasy chemie genetika metabolismus MeSH
- výpočetní biologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
