A method has been developed that automatically fits double-helical regions into the electron density of nucleic acid structures. Rigid fragments consisting of two Watson-Crick base pairs and three pairs of phosphate groups in the A-type or B-type conformation are positioned into the electron density by phased rotation and translation functions. The position and orientation of the localized double-helical fragments are determined by phased refinement. The method has been tested by building double-helical regions of nine RNA structures of variable crystallographic resolution and polynucleotide length and is available for free use.

• MeSH
• DNA chemie   MeSH
• financování organizované MeSH
• konformace nukleové kyseliny MeSH
• krystalografie rentgenová MeSH
• molekulární modely MeSH
• párování bází MeSH
• počítačová simulace MeSH
• RNA chemie   MeSH
• software MeSH
• statická elektřina MeSH
• Publikační typ
• srovnávací studie MeSH
• validační studie MeSH

Metabolic flux investigations of cells and tissue samples are a rapidly advancing tool in diverse research areas. Reliable methods of data normalization are crucial for an adequate interpretation of results and to avoid a misinterpretation of experiments and incorrect conclusions. The most common methods for metabolic flux data normalization are to cell number, DNA and protein. Data normalization may be affected by a variety of factors, such as density, healthy state, adherence efficiency, or proportional seeding of cells. The mussel-derived adhesive Cell-Tak is often used to immobilize poorly adherent cells. Here we demonstrate that this coating strongly affects the fluorescent detection of DNA leading to an incorrect and highly variable normalization of metabolic flux data. Protein assays are much less affected and cell counting can virtually completely remove the effect of the coating. Cell-Tak coating also affects cell shape in a cell line-specific manner and may change cellular metabolism. Based on these observations we recommend cell counting as a gold standard normalization method for Seahorse metabolic flux measurements with protein content as a reasonable alternative.

• MeSH
• DNA * MeSH
• membránové proteiny * MeSH
• Publikační typ
• časopisecké články MeSH

The sperm of sterlet (Acispenser ruthenus) was used to investigate the effect of the xenobiotic tetrabrombisphenol A (TBBPA) on sperm quality variables (ATP content, spermatozoa motility, and velocity), DNA integrity, and oxidative stress indices. Sperm was diluted to obtain a spermatozoa density of 5 × 10(8) cells/mL and exposed for 2 h to final concentrations of TBBPA (0.5, 1.75, 2.5, 5, and 10 μg/L). The oxidative stress indices, including lipid peroxidation, carbonyl derivatives of proteins, and antioxidant activity were significantly higher with increased concentrations of TBBPA. There was significantly less intracellular ATP in sperm samples at TBBPA concentrations of 2.5 μg/L and above. Spermatozoa velocity and percent motile sperm were significantly lower at each sampling time post-activation compared to controls. DNA damage expressed as percent DNA in Tail and Olive Tail moment was significantly higher with exposures ≥2.5 μg/L TBBPA. The results demonstrated that TBBPA and other xenobiotics can induce reactive oxygen species stress in fish spermatozoa, which could impair the sperm quality, DNA integrity, ATP content, and the antioxidant defense system. This study confirmed that fish spermatozoa can be used in in vitro assays for monitoring residual pollution in aquatic environments.

• MeSH
• adenosintrifosfát metabolismus   MeSH
• antioxidancia metabolismus   MeSH
• benzhydrylové sloučeniny chemie   toxicita   MeSH
• DNA chemie   metabolismus   MeSH
• fenoly chemie   toxicita   MeSH
• karbonylace proteinů účinky léků   MeSH
• motilita spermií účinky léků   MeSH
• oxidační stres účinky léků   MeSH
• peroxidace lipidů účinky léků   MeSH
• polybrombifenylové sloučeniny chemie   toxicita   MeSH
• poškození DNA účinky léků   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• ryby metabolismus   MeSH
• spermie fyziologie   MeSH
• superoxiddismutasa metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Although oocyst morphology was always considered as a reliable parameter for coccidian species discrimination we describe strain variation of turkey coccidia, Eimeria adenoeides, which remarkably exceeds the variation observed in any other Eimeria species. Two strains have been isolated - the first strain maintains the typical oocyst morphology attributed to this species - large and ellipsoidal - while the second strain has small and ovoid oocysts, never described before for this species. Other biological parameters including pathogenicity were found to be similar. Cross-protection between these 2 strains in 2 immunization and challenge experiments was confirmed. Sequencing and analysis of 18S and ITS1 ribosomal DNA revealed a close relationship according to 18S and a relatively distant relationship according to ITS1. Analysis of 18S and ITS1 sequences from commercial turkey coccidiosis vaccines Immucox®-T and Coccivac®-T revealed that each vaccine contains a different strain of E. adenoeides and that these strains have 18S and ITS1 sequences homologous to the sequences of the strains we have isolated and described. These findings show that diagnostics of turkey coccidia according to oocyst morphology have to be carried out with caution or abolished entirely. Novel PCR-based molecular tools will be necessary for fast and reliable species discrimination.

• MeSH
• DNA vakcíny aplikace a dávkování   MeSH
• druhová specificita MeSH
• Eimeria cytologie   genetika   izolace a purifikace   patogenita   MeSH
• feces parazitologie   MeSH
• fylogeneze MeSH
• genetická variace MeSH
• kokcidióza diagnóza   imunologie   parazitologie   prevence a kontrola   MeSH
• krocani imunologie   parazitologie   MeSH
• mikroskopie MeSH
• molekulární typizace MeSH
• nemoci drůbeže diagnóza   imunologie   parazitologie   prevence a kontrola   MeSH
• oocysty cytologie   MeSH
• počet buněk MeSH
• polymerázová řetězová reakce MeSH
• respirační komplex IV analýza   MeSH
• RNA ribozomální 18S analýza   MeSH
• sekvenční analýza DNA MeSH
• vakcinace MeSH
• zkřížená ochrana účinky léků   imunologie   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Establishing translocated populations is a common process to preserve and maintain genetic diversity of threatened species. In 2001, three translocated populations of noble crayfish (Astacus astacus) were established in the Czech Republic, founded by either adult or juvenile individuals from three particular source populations. We assessed genetic diversity at seven microsatellite loci after one decade (assumed three generations) from establishment. Although the translocated populations exhibited a slight but non-significant reduction in genetic diversity (A R = 2.2-5.0; H O = 0.11-0.31), the most striking result was generally very low genetic diversity in source populations (A R = 3.0-5.3; H O = 0.15-0.38). Similarly, a high degree of inbreeding (F IS = 0.36-0.60) demonstrates the nature of source populations, already affected by isolation and small size. In spite of that, based on the results of this study, the establishment of new translocated noble crayfish populations was successful, since there is no significant decline in genetic variability and all populations are still viable. Although source populations did not exhibit high genetic diversity, their distinctiveness makes them possible to use for conservation purposes. Continued monitoring is necessary to track the long-term progress of the translocation program, including other parameters describing the state of the population, such as the occurrence and frequency of diseases or morphological changes.

• MeSH
• alely MeSH
• efekt zakladatele * MeSH
• genetická variace * MeSH
• hustota populace MeSH
• inbreeding MeSH
• mikrosatelitní repetice MeSH
• populační genetika * MeSH
• sekvenční analýza DNA MeSH
• severní raci genetika   MeSH
• zachování přírodních zdrojů MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

Coccidian oocysts from feces of 46 individuals of the garden dormouse, Eliomys quercinus (Rodentia: Gliridae), were morphologically and molecularly characterized. Both morphological and sequence data (18S rDNA and ORF 470) showed low variability, indicating that all samples represent a single species. By comparison with published morphological descriptions of coccidia from glirid rodents, we determined that the samples represent Eimeria myoxi. Molecular data suggest that this species does not fall within the 2 known rodent-specific groups but branches as a third independent lineage. However, its exact position in respect to other eimerian clusters could not be established due to the lack of phylogenetic information at this taxonomic level for the 18S rRNA and ORF 470 genes. Based on these results, we provide a re-description of Eimeria myoxi, which contains morphological and molecular characteristics sufficient for its further unequivocal identification.

• MeSH
• Eimeria cytologie   genetika   izolace a purifikace   MeSH
• feces parazitologie   MeSH
• fylogeneze MeSH
• kokcidióza diagnóza   parazitologie   MeSH
• mikroskopie MeSH
• molekulární typizace MeSH
• Myoxidae parazitologie   MeSH
• nemoci hlodavců diagnóza   parazitologie   MeSH
• oocysty cytologie   MeSH
• počet buněk MeSH
• polymerázová řetězová reakce MeSH
• protozoální DNA analýza   MeSH
• respirační komplex IV analýza   MeSH
• RNA ribozomální 18S analýza   MeSH
• sekvenční analýza DNA MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: TDP-43 proteinopathies represent a spectrum of neurodegenerative disorders. Variable clinical presentations including frontotemporal dementia, amyotrophic lateral sclerosis (ALS) and mixed forms are associated with the spatial heterogeneity of the TDP-43 pathology. Recent studies have emphasized the role of oligodendrocytes in the pathogenesis of ALS. OBJECTIVE: To evaluate whether TDP-43 proteinopathies are associated with an oligodendroglial response. METHODS: We performed a study on 7 controls and 10 diseased patients with spinal cord involvement. Using the oligodendroglia-specific antibody TPPP/p25, we assessed oligodendrocyte density in the lateral corticospinal tracts (LCSs) along with the presence of perineuronal oligodendrocytes (PNOGs) in the anterior horns. We performed a densitometry of myelin basic protein (MBP) immunoreactivity. The numbers of TDP-43 and p62 immunoreactive inclusions were counted in both the LCSs and the anterior horns. RESULTS: Double immunolabeling confirmed that oligodendrocytes harbor TDP-43 inclusions. In the LCSs, MBP density, but not the number of oligodendrocytes, was decreased in the diseased group. However, oligodendrocyte counts in the LCS correlated positively, and the density of MBP inversely, with the number of neuronal inclusions in the anterior horn, suggestive of a compensatory response of oligodendrocytes. The number of neurons with PNOGs correlated with the amount of inclusions. CONCLUSION: Our study further emphasizes the importance of oligodendroglia in the pathogenesis of TDP-43 proteinopathies with spinal cord involvement.

• MeSH
• buněčná inkluze metabolismus   patologie   MeSH
• buňky předních rohů míšních metabolismus   patologie   MeSH
• DNA vazebné proteiny metabolismus   MeSH
• dospělí MeSH
• encefalitogenní základní proteiny metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mícha metabolismus   patologie   MeSH
• mladý dospělý MeSH
• motorické neurony metabolismus   patologie   MeSH
• oligodendroglie metabolismus   patologie   MeSH
• proteinopatie TDP-43 metabolismus   patologie   MeSH
• proteiny vázající RNA metabolismus   MeSH
• pyramidové dráhy metabolismus   patologie   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Tautomerism of nucleic acid (NA) bases is a crucial factor for the maintenance and translation of genetic information in organisms. Only canonical tautomers of NA bases can form hydrogen-bonded complexes with their natural counterparts. On the other hand, rare tautomers of nucleobases have been proposed to be involved in processes catalysed by NA enzymes. Isocytosine, which can be considered as a structural fragment of guanine, is known to have two stable tautomers both in solution and solid states. The tautomer equilibrium of isocytosine contrasts with the remarkable stability of the canonical tautomer of guanine. This paper investigates the factors contributing to the stability of the canonical tautomer of guanine by a combination of NMR experiments and theoretical calculations. The electronic effects of substituents on the stability of the rare tautomers of isocytosine and guanine derivatives are studied by density functional theory (DFT) calculations. Selected derivatives are studied by variable-temperature NMR spectroscopy. Rare tautomers can be stabilised in solution by intermolecular hydrogen-bonding interactions with suitable partners. These intermolecular interactions give rise to characteristic signals in proton NMR spectra, which make it possible to undoubtedly confirm the presence of a rare tautomer.

• MeSH
• cytosin analogy a deriváty   chemie   MeSH
• dimerizace MeSH
• DNA chemie   MeSH
• elektrony MeSH
• guanin chemie   MeSH
• kvantová teorie MeSH
• magnetická rezonanční spektroskopie metody   MeSH
• makromolekulární látky MeSH
• normální rozdělení MeSH
• reprodukovatelnost výsledků MeSH
• stereoizomerie MeSH
• termodynamika MeSH
• vodík chemie   MeSH
• vodíková vazba MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Osteoporóza je z 60- 80 % onemocněním dědičným s charakteristickou multifaktoriální patogenezí, v níž dochází k interakci vlivu mnoha "slabých" genů a vnějších faktorů. Zatím nejvíce informací o vztahu genů k variabilitě kostních parametrů (denzitě, kvalitě a metabolizmu) přinesly asociační studie kandidátních genů pro osteoporózu. Z nejznámějších genů vztahujících se ke kostní denzitě byly identifikovány geny pro receptor vitaminu D, estrogen a kalcitonin, LRP5 a LRP6. Za parametry kostní remodelace jsou odpovědné geny pro IL-1a a osteoprotegerin. Z nedávno objevených genů vztahujících se ke kostnímu fenotypu byly identifikovány geny pro hypolaktázii, tetrafolát reduktázu a ALDH7A1. Rozměry kosti pravděpodobně částečně kontroluje gen PLCL1. Kandidátní geny pro osteoporózu pravděpodobně determinují i produkci kalciotropních hormonů (PTH, sexuální steroidy), a dokonce i některé mimokostní fenotypy (zánět, imunita, sklon k malignitám). Naopak geny determinující mimokostní parametry (např. hladiny lipoproteinů) jsou asociovány s fenotypem kostním (gen pro ApoE se vztahuje ke kostní denzitě). Asociační studie však mají závažná omezení, mj. také vlivem vazebné nerovnováhy související s těsnou blízkostí identifikovaných genů v DNK, jež může být jednou z příčin falešně pozitivních výsledků. U dětí, kde budování skeletu ovlivňují především faktory vnější (nutrice, pohybová aktivita), je vztah kandidátních genů ke kostní hmotě méně těsný než u dospělých. Přehledná práce se zabývá fyziologií a sexuální diferenciací pubertální kosti. Diskutován je význam identifikace kandidátních genů pro prevenci a cílenou léčbu osteoporózy (farmakogenetika) a využití programu FRAX (WHO) pro desetiletou predikci fraktur osteopenických pacientů.

Osteoporosis is in 60–?80% a hereditary disease with a characteristic multifactorial pathogenesis during which the effects of many “weak” genes interact with external factors. To date, most information relating to the correlations between genes and bone parameter variability (density, quality and metabolism) has been provided by association studies of candidate genes for osteoporosis. The best known genes related to bone density have been identified as the genes for the vitamin D, estrogen and calcitonin receptor, LRP5 and LRP6. The genes for IL-1? and osteoprotegerin are responsible for the parameters of bone remodeling. Recently discovered genes related to bone phenotype include identified genes for hypolactasia, tetrafolate reductase and ALDH7A1. Bone size and dimensions are probably partially controlled by the PLCL1 gene. Candidate genes for osteoporosis probably also determine the production of calciotropic hormones (PTH, sex steroids) and even some extra-osseous phenotypes (inflammation, immunity, susceptibility to malignancies). On the contrary, genes that determine extra-osseous parameters (e.?g. lipoprotein levels) are associated with the bone phenotype (the gene for ApoE is related to bone density). Association studies, though, have serious limitations. Among others, these include the influence of linkage disequilibrium associated with the close proximity of the identified genes within DNA, which may be one of the causes of false positive results. In children, where building of the skeleton is influenced predominantly by external factors (nutrition, physical activity), the relationship between candidate genes and bone mass is less close than in adults. This overview deals with the physiology and sexual differentiation of pubertal bone. It discusses the importance of identifying candidate genes in the prevention and targeted treatment of osteoporosis (pharmacogenetics) as well as the application of the FRAX (WHO) program in the ten-year prediction of fractures in osteopenic patients.

• Klíčová slova
• kandidátní geny pro osteoporózu, pubertální kost, prevence osteoporózy,
• MeSH
• celogenomová asociační studie MeSH
• farmakogenetika MeSH
• genetická predispozice k nemoci MeSH
• kostní denzita genetika   MeSH
• lidé MeSH
• osteoporóza diagnóza   etiologie   genetika   patofyziologie   MeSH
• remodelace kosti genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• přehledy MeSH

BACKGROUND: QTL cloning for the discovery of genes underlying polygenic traits has historically been cumbersome in long-lived perennial plants like Populus. Linkage disequilibrium-based association mapping has been proposed as a cloning tool, and recent advances in high-throughput genotyping and whole-genome resequencing enable marker saturation to levels sufficient for association mapping with no a priori candidate gene selection. Here, multiyear and multienvironment evaluation of cell wall phenotypes was conducted in an interspecific P. trichocarpa x P. deltoides pseudo-backcross mapping pedigree and two partially overlapping populations of unrelated P. trichocarpa genotypes using pyrolysis molecular beam mass spectrometry, saccharification, and/ or traditional wet chemistry. QTL mapping was conducted using a high-density genetic map with 3,568 SNP markers. As a fine-mapping approach, chromosome-wide association mapping targeting a QTL hot-spot on linkage group XIV was performed in the two P. trichocarpa populations. Both populations were genotyped using the 34 K Populus Infinium SNP array and whole-genome resequencing of one of the populations facilitated marker-saturation of candidate intervals for gene identification. RESULTS: Five QTLs ranging in size from 0.6 to 1.8 Mb were mapped on linkage group XIV for lignin content, syringyl to guaiacyl (S/G) ratio, 5- and 6-carbon sugars using the mapping pedigree. Six candidate loci exhibiting significant associations with phenotypes were identified within QTL intervals. These associations were reproducible across multiple environments, two independent genotyping platforms, and different plant growth stages. cDNA sequencing for allelic variants of three of the six loci identified polymorphisms leading to variable length poly glutamine (PolyQ) stretch in a transcription factor annotated as an ANGUSTIFOLIA C-terminus Binding Protein (CtBP) and premature stop codons in a KANADI transcription factor as well as a protein kinase. Results from protoplast transient expression assays suggested that each of the polymorphisms conferred allelic differences in the activation of cellulose, hemicelluloses, and lignin pathway marker genes. CONCLUSION: This study illustrates the utility of complementary QTL and association mapping as tools for gene discovery with no a priori candidate gene selection. This proof of concept in a perennial organism opens up opportunities for discovery of novel genetic determinants of economically important but complex traits in plants.

• MeSH
• alely MeSH
• buněčná stěna genetika   MeSH
• celulosa metabolismus   MeSH
• fenotyp MeSH
• genetická vazba MeSH
• genotyp MeSH
• jednonukleotidový polymorfismus MeSH
• lignin biosyntéza   MeSH
• lod skóre MeSH
• lokus kvantitativního znaku MeSH
• mapování chromozomů MeSH
• Populus genetika   MeSH
• rostlinné geny * MeSH
• rostlinné proteiny chemie   genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční seřazení MeSH
• transkripční faktory chemie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH