Crop inoculation with Glomus cubense isolate (INCAM-4, DAOM-241198) promotes yield in banana, cassava, forages, and others. Yield improvements range from 20 to 80% depending on crops, nutrient supply, and edaphoclimatic conditions. However, it is difficult to connect yield effects with G. cubense abundance in roots due to the lack of an adequate methodology to trace this taxon in the field. It is necessary to establish an accurate evaluation framework of its contribution to root colonization separated from native arbuscular mycorrhizal fungi (AMF). A taxon-discriminating primer set was designed based on the ITS nrDNA marker and two molecular approaches were optimized and validated (endpoint PCR and quantitative real-time PCR) to trace and quantify the G. cubense isolate in root and soil samples under greenhouse and environmental conditions. The detection limit and specificity assays were performed by both approaches. Different 18 AMF taxa were used for endpoint PCR specificity assay, showing that primers specifically amplified the INCAM-4 isolate yielding a 370 bp-PCR product. In the greenhouse, Urochloa brizantha plants inoculated with three isolates (Rhizophagus irregularis, R. clarus, and G. cubense) and environmental root and soil samples were successfully traced and quantified by qPCR. The AMF root colonization reached 41-70% and the spore number 4-128 per g of soil. This study demonstrates for the first time the feasibility to trace and quantify the G. cubense isolate using a taxon-discriminating ITS marker in roots and soils. The validated approaches reveal their potential to be used for the quality control of other mycorrhizal inoculants and their relative quantification in agroecosystems.
- MeSH
- genetické markery * genetika MeSH
- Glomeromycota genetika MeSH
- houby genetika MeSH
- kořeny rostlin mikrobiologie MeSH
- lipnicovité mikrobiologie MeSH
- mykorhiza * genetika MeSH
- polymerázová řetězová reakce MeSH
- půdní mikrobiologie * MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- genetické markery * MeSH
Arabidopsis thaliana 45S ribosomal genes (rDNA) are located in tandem arrays called nucleolus organizing regions on the termini of chromosomes 2 and 4 (NOR2 and NOR4) and encode rRNA, a crucial structural element of the ribosome. The current model of rDNA organization suggests that inactive rRNA genes accumulate in the condensed chromocenters in the nucleus and at the nucleolar periphery, while the nucleolus delineates active genes. We challenge the perspective that all intranucleolar rDNA is active by showing that a subset of nucleolar rDNA assembles into condensed foci marked by H3.1 and H3.3 histones that also contain the repressive H3K9me2 histone mark. By using plant lines containing a low number of rDNA copies, we further found that the condensed foci relate to the folding of rDNA, which appears to be a common mechanism of rDNA regulation inside the nucleolus. The H3K9me2 histone mark found in condensed foci represents a typical modification of bulk inactive rDNA, as we show by genome-wide approaches, similar to the H2A.W histone variant. The euchromatin histone marks H3K27me3 and H3K4me3, in contrast, do not colocalize with nucleolar foci and their overall levels in the nucleolus are very low. We further demonstrate that the rDNA promoter is an important regulatory region of the rDNA, where the distribution of histone variants and histone modifications are modulated in response to rDNA activity.
- Klíčová slova
- chromatin, chromatin immunoprecipitation, histone variants, nucleolar foci, rDNA, super-resolution microscopy,
- MeSH
- Arabidopsis genetika MeSH
- buněčné jadérko genetika MeSH
- buněčné jádro genetika MeSH
- DNA rostlinná genetika metabolismus MeSH
- epigeneze genetická genetika MeSH
- genetická transkripce MeSH
- genetická variace MeSH
- genetické markery genetika MeSH
- histony genetika metabolismus MeSH
- kořeny rostlin metabolismus MeSH
- ribozomální DNA genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA rostlinná MeSH
- genetické markery MeSH
- histony MeSH
- ribozomální DNA MeSH
Though apple genotyping is mainly used for scientific and breeding purposes, it can also be adopted by national authorities to control the authenticity of apple cultivars. To facilitate the introduction of routine apple genotyping into practice, a new apple simple sequence repeat (SSR) genotyping kit was developed (called the Ap17 in. SSR Genotyping Kit). The kit combines 17 SSR markers including those recommended by the Working Group of the European Cooperative Programme for Plant Genetic Resources (ECPGR), covering all apple linkage groups in a one-tube reaction format, using a fragment analysis method to simplify the genotyping procedure. The kit was successfully tested using 880 unique diploid apple germplasm accessions; the kit can also readily discriminate triploid and tetraploid samples. The total probability of identity for the kit and the sample collection used was calculated to be 1.73 × 10-22. Tables for converting results to enable genotype comparisons between currently-used genotyping systems and the Ap17 in. kit are provided. The kit is ideally suited for validation in laboratories genotyping a large number of apple samples, saving time, costs, and labor, while minimizing technical and human errors.
- Klíčová slova
- Apple, Genotyping, Multiplex PCR, SSR, Simple sequence repeat,
- MeSH
- diploidie MeSH
- genetické markery genetika MeSH
- genotypizační techniky metody MeSH
- Malus genetika MeSH
- mikrosatelitní repetice genetika MeSH
- multiplexová polymerázová řetězová reakce MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- genetické markery MeSH
Prevalence of inflammatory bowel disease (IBD), a chronic inflammatory disorder of the gut, has been on the rise in recent years-not only in the adult population but also especially in pediatric patients. Despite the absence of curative treatments, current therapeutic options are able to achieve long-term remission in a significant proportion of cases. To this end, however, there is a need for biomarkers enabling accurate diagnosis, prognosis, and prediction of response to therapies to facilitate a more individualized approach to pediatric IBD patients. In recent years, evidence has continued to evolve concerning noncoding RNAs (ncRNAs) and their roles as integral factors in key immune-related cellular pathways. Specific deregulation patterns of ncRNAs have been linked to pathogenesis of various diseases, including pediatric IBD. In this article, we provide an overview of current knowledge on ncRNAs, their altered expression profiles in pediatric IBD patients, and how these are emerging as potentially valuable clinical biomarkers as we enter an era of personalized medicine.
- Klíčová slova
- Crohn’s disease, inflammatory bowel disease, microRNA, noncoding RNA, pediatrics, ulcerative colitis,
- MeSH
- biologické markery analýza MeSH
- Crohnova nemoc genetika MeSH
- dítě MeSH
- genetické markery genetika MeSH
- idiopatické střevní záněty genetika MeSH
- individualizovaná medicína trendy MeSH
- lidé MeSH
- nekódující RNA analýza MeSH
- signální transdukce genetika MeSH
- transkriptom MeSH
- ulcerózní kolitida genetika MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- úvodní články MeSH
- Názvy látek
- biologické markery MeSH
- genetické markery MeSH
- nekódující RNA MeSH
We sought to analyse plasma levels of peripheral blood microRNAs (miRs) as biomarkers of ST-segment-elevation myocardial infarction (STEMI) due to type-1 myocardial infarction as a model situation of vulnerable plaque (VP) rupture. Samples of 20 patients with STEMI were compared both with a group of patients without angina pectoris in whom coronary angiogram did not reveal coronary atherosclerotic disease (no coronary atherosclerosis-NCA) and a group of patients with stable angina pectoris and at least one significant coronary artery stenosis (stable coronary artery disease-SCAD). This study design allowed us to identify miRs deregulated in the setting of acute coronary artery occlusion due to VP rupture. Based on an initial large scale miR assay screening, we selected a total of 12 miRs (three study miRs and nine controls) that were tested in the study. Two of the study miRs (miR-331 and miR-151-3p) significantly distinguished STEMI patients from the control groups, while ROC analysis confirmed their suitability as biomarkers. Importantly, this was observed in patients presenting early with STEMI, even before the markers of myocardial necrosis (cardiac troponin I, miR-208 and miR-499) were elevated, which suggests that the origin of miR-331 and miR-151-3p might be in the VP. In conclusion, the study provides two novel biomarkers observed in STEMI, which may be associated with plaque rupture.
- MeSH
- akutní koronární syndrom genetika MeSH
- genetické markery genetika MeSH
- infarkt myokardu s elevacemi ST úseků genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- mikro RNA genetika MeSH
- nemoci koronárních tepen genetika MeSH
- senioři MeSH
- studie případů a kontrol MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- genetické markery MeSH
- mikro RNA MeSH
- MIRN151a microRNA, human MeSH Prohlížeč
- MIRN331 microRNA, human MeSH Prohlížeč
Analysis of population genetic structure has become a standard approach in population genetics. In polyploid complexes, clustering analyses can elucidate the origin of polyploid populations and patterns of admixture between different cytotypes. However, combining diploid and polyploid data can theoretically lead to biased inference with (artefactual) clustering by ploidy. We used simulated mixed-ploidy (diploid-autotetraploid) data to systematically compare the performance of k-means clustering and the model-based clustering methods implemented in STRUCTURE, ADMIXTURE, FASTSTRUCTURE and INSTRUCT under different scenarios of differentiation and with different marker types. Under scenarios of strong population differentiation, the tested applications performed equally well. However, when population differentiation was weak, STRUCTURE was the only method that allowed unbiased inference with markers with limited genotypic information (co-dominant markers with unknown dosage or dominant markers). Still, since STRUCTURE was comparatively slow, the much faster but less powerful FASTSTRUCTURE provides a reasonable alternative for large datasets. Finally, although bias makes k-means clustering unsuitable for markers with incomplete genotype information, for large numbers of loci (>1000) with known dosage k-means clustering was superior to FASTSTRUCTURE in terms of power and speed. We conclude that STRUCTURE is the most robust method for the analysis of genetic structure in mixed-ploidy populations, although alternative methods should be considered under some specific conditions.
- MeSH
- diploidie MeSH
- genetická variace genetika MeSH
- genetické markery genetika MeSH
- genotyp MeSH
- jednonukleotidový polymorfismus genetika MeSH
- mikrosatelitní repetice genetika MeSH
- ploidie * MeSH
- populační genetika statistika a číselné údaje MeSH
- shluková analýza MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- genetické markery MeSH
Amplicon-based next-generation sequencing (NGS) of immunoglobulin (IG) and T-cell receptor (TR) gene rearrangements for clonality assessment, marker identification and quantification of minimal residual disease (MRD) in lymphoid neoplasms has been the focus of intense research, development and application. However, standardization and validation in a scientifically controlled multicentre setting is still lacking. Therefore, IG/TR assay development and design, including bioinformatics, was performed within the EuroClonality-NGS working group and validated for MRD marker identification in acute lymphoblastic leukaemia (ALL). Five EuroMRD ALL reference laboratories performed IG/TR NGS in 50 diagnostic ALL samples, and compared results with those generated through routine IG/TR Sanger sequencing. A central polytarget quality control (cPT-QC) was used to monitor primer performance, and a central in-tube quality control (cIT-QC) was spiked into each sample as a library-specific quality control and calibrator. NGS identified 259 (average 5.2/sample, range 0-14) clonal sequences vs. Sanger-sequencing 248 (average 5.0/sample, range 0-14). NGS primers covered possible IG/TR rearrangement types more completely compared with local multiplex PCR sets and enabled sequencing of bi-allelic rearrangements and weak PCR products. The cPT-QC showed high reproducibility across all laboratories. These validated and reproducible quality-controlled EuroClonality-NGS assays can be used for standardized NGS-based identification of IG/TR markers in lymphoid malignancies.
- MeSH
- akutní lymfatická leukemie genetika MeSH
- genetické markery genetika MeSH
- genová přestavba T-lymfocytů genetika MeSH
- geny pro imunoglobuliny genetika MeSH
- geny TcR genetika MeSH
- imunoglobuliny genetika MeSH
- lidé MeSH
- receptory antigenů T-buněk genetika MeSH
- referenční standardy MeSH
- rekombinace genetická genetika MeSH
- reprodukovatelnost výsledků MeSH
- reziduální nádor genetika MeSH
- výpočetní biologie metody MeSH
- vysoce účinné nukleotidové sekvenování metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- genetické markery MeSH
- imunoglobuliny MeSH
- receptory antigenů T-buněk MeSH
Assessment of clonality, marker identification and measurement of minimal residual disease (MRD) of immunoglobulin (IG) and T cell receptor (TR) gene rearrangements in lymphoid neoplasms using next-generation sequencing (NGS) is currently under intensive development for use in clinical diagnostics. So far, however, there is a lack of suitable quality control (QC) options with regard to standardisation and quality metrics to ensure robust clinical application of such approaches. The EuroClonality-NGS Working Group has therefore established two types of QCs to accompany the NGS-based IG/TR assays. First, a central polytarget QC (cPT-QC) is used to monitor the primer performance of each of the EuroClonality multiplex NGS assays; second, a standardised human cell line-based DNA control is spiked into each patient DNA sample to work as a central in-tube QC and calibrator for MRD quantification (cIT-QC). Having integrated those two reference standards in the ARResT/Interrogate bioinformatic platform, EuroClonality-NGS provides a complete protocol for standardised IG/TR gene rearrangement analysis by NGS with high reproducibility, accuracy and precision for valid marker identification and quantification in diagnostics of lymphoid malignancies.
- MeSH
- genetické markery genetika MeSH
- genová přestavba genetika MeSH
- imunoglobuliny genetika MeSH
- lidé MeSH
- receptory antigenů T-buněk genetika MeSH
- reprodukovatelnost výsledků MeSH
- reziduální nádor genetika MeSH
- řízení kvality MeSH
- výpočetní biologie metody MeSH
- vysoce účinné nukleotidové sekvenování metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- genetické markery MeSH
- imunoglobuliny MeSH
- receptory antigenů T-buněk MeSH
West Nile virus (WNV) is a mosquito-borne zoonotic neurotropic virus capable to cause lethal meningoencephalitis (WNE) in infected hosts such as birds, horses, and humans. Due to their sensitivity, horses serve as sentinel species in areas at risk. We studied a population of Camargue horses living in Southern France in two zones with endemic WNV circulation where WNV outbreaks were recorded in 2000 and 2003-4. Two sets of microsatellite markers located in MHC and Ly49 genomic regions were genotyped as well as multiple SNPs in ten immunity-related candidate gene regions. Associations between genetic polymorphisms and resistance/susceptibility to WNE were tested. While single marker associations were weak, compound two-gene genotypes of SNPs located within the MAVS, NCR2 and IL-10 genes and microsatellites HMS082 and CZM013 were associated with susceptibility to WNE. Combinations of microsatellite markers CZM009, ABGe17402 and ABGe9019 were associated with simple seroconversion without clinical signs of WNE (resistance). In addition, a distribution of polymorphic markers between WNV-IgG seropositive horses and a control group of WNV-IgG seronegative horses was tested. One SNP in the OAS1 gene (NC_009151.3:g.21961328A>G) was significantly associated with the seropositive phenotype (pcorr = 0.023; OR = 40.5 CI (4.28; 383.26); RR = 8.18 CI (1.27; 52.89) in the Camargue breed. In compound genotypes, SNP markers for SLC11A1, MAVS, OAS1, TLR4, ADAM17 and NCR2 genes and ten microsatellites showed non-random distribution between seropositive and seronegative groups of horses. Further analysis of associated markers could contribute to our understanding of anti-WNV defense mechanisms in horses.
- Klíčová slova
- Associations, Camargue, Horse, Immunity-related candidate gene SNP markers, MHC and Ly49 microsatellite markers, West Nile virus,
- MeSH
- genetická predispozice k nemoci genetika MeSH
- genetické markery genetika MeSH
- imunoglobulin G imunologie MeSH
- jednonukleotidový polymorfismus MeSH
- koně MeSH
- mikrosatelitní repetice MeSH
- nemoci koní genetika virologie MeSH
- odolnost vůči nemocem genetika MeSH
- polymorfismus genetický * MeSH
- virus západního Nilu fyziologie MeSH
- západonilská horečka genetika veterinární virologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Francie epidemiologie MeSH
- Názvy látek
- genetické markery MeSH
- imunoglobulin G MeSH
MOTIVATION: Modern molecular methods have increased our ability to describe microbial communities. Along with the advances brought by new sequencing technologies, we now require intensive computational resources to make sense of the large numbers of sequences continuously produced. The software developed by the scientific community to address this demand, although very useful, require experience of the command-line environment, extensive training and have steep learning curves, limiting their use. We created SEED 2, a graphical user interface for handling high-throughput amplicon-sequencing data under Windows operating systems. RESULTS: SEED 2 is the only sequence visualizer that empowers users with tools to handle amplicon-sequencing data of microbial community markers. It is suitable for any marker genes sequences obtained through Illumina, IonTorrent or Sanger sequencing. SEED 2 allows the user to process raw sequencing data, identify specific taxa, produce of OTU-tables, create sequence alignments and construct phylogenetic trees. Standard dual core laptops with 8 GB of RAM can handle ca. 8 million of Illumina PE 300 bp sequences, ca. 4 GB of data. AVAILABILITY AND IMPLEMENTATION: SEED 2 was implemented in Object Pascal and uses internal functions and external software for amplicon data processing. SEED 2 is a freeware software, available at http://www.biomed.cas.cz/mbu/lbwrf/seed/ as a self-contained file, including all the dependencies, and does not require installation. Supplementary data contain a comprehensive list of supported functions. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
