FISH-aimed karyotype analysis in Aconitum subgen : aconitum reveals excessive rDNA sites in tetraploid taxa

The location of 5S and 35S rDNA sequences in chromosomes of four Aconitum subsp. Aconitum species was analyzed after fluorescence in situ hybridization (FISH). Both in diploids (2n = 2x = 16; Aconitum variegatum, A. degenii) and tetraploids (2n = 4× = 32; A. firmum, A. plicatum), rDNA repeats were localized exclusively on the shorter arms of chromosomes, in subterminal or pericentromeric sites. All analyzed species showed similar basal genome size (Cx = 5.31-5.71 pg). The most striking features of tetraploid karyotypes were the conservation of diploid rDNA loci and emergence of many additional 5S rDNA clusters. Chromosomal distribution of excessive ribosomal sites suggests their role in the secondary diploidization of tetraploid karyotypes

On the radiogenic heat production of igneous rocks

Radiogenic heat production is a physical parameter crucial to properly estimating lithospheric temperatures and properly understanding processes related to the thermal evolution of the Earth. Yet heat production is, in general, poorly constrained by direct observation because the key radiogenic elements exist in trace amounts making them difficulty image geophysically. In this study, we advance our knowledge of heat production throughout the lithosphere by analyzing chemical analyses of 108,103 igneous rocks provided by a number of geochemical databases. We produce global estimates of the average and natural range for igneous rocks using common chemical classification systems. Heat production increases as a function of increasing felsic and alkali content with similar values for analogous plutonic and volcanic rocks. The logarithm of median heat production is negatively correlated (r2 ¼ 0.98) to compositionally-based estimates of seismic velocities between 6.0 and 7.4 km s 1, consistent with the vast majority of igneous rock compositions. Compositional variations for continent-wide models are also well-described by a log-linear correlation between heat production and seismic velocity. However, there are differences between the log-linear models for North America and Australia, that are consistent with interpretations from previous studies that suggest above average heat production across much of Australia. Similar log-linear models also perform well within individual geological provinces withw1000 samples. This correlation raises the prospect that this empirical method can be used to estimate average heat production and natural variance both laterally and vertically throughout the lithosphere. This correlative relationship occurs despite a direct causal relationship between these two parameters but probably arises from the process of differentiation through melting and crystallization.D. Hasterok, J. Web

Epigenetics of the preferential silencing of Brachypodium stacei-originated 35S rDNA loci in the allotetraploid grass Brachypodium hybridum

Nucleolar dominance (ND), initially described as 'differential amphiplasty', is a phenomenon observed in some plant and animal allopolyploids and hybrids in which the selective suppression of the activity of 35S rRNA gene loci that have been inherited from one of the two or more ancestral genomes occurs. Although more than 80 years have passed since the discovery of ND, there is still a significant lack in our understanding of the mechanisms that determine this phenomenon. Here, we aimed to investigate the epigenetic status of 35S rRNA gene loci in the monocotyledonous Brachypodium hybridum, which is an allotetraploid that has resulted from a cross between B. distachyon and B. stacei. We revealed that the repressed B. stacei-inherited rDNA loci are characterised by a high level of DNA methylation. The global hypomethylation of B. hybridum nuclear DNA induced by 5-azacytidine, however, seems to be insufficient for the transcriptional reactivation of these loci, which indicates that factors other than DNA methylation are behind the suppression of B. stacei-originated loci. We also showed that the transcriptionally active and silenced fractions of rRNA genes that had been inherited from B. distachyon occupy different domains within the chromocentres adjacent to the nucleolus, depending on their epigenetic status

Reconstructing the evolution of Brachypodium genomes using comparative chromosome painting.

Brachypodium distachyon is a model for the temperate cereals and grasses and has a biology, genomics infrastructure and cytogenetic platform fit for purpose. It is a member of a genus with fewer than 20 species, which have different genome sizes, basic chromosome numbers and ploidy levels. The phylogeny and interspecific relationships of this group have not to date been resolved by sequence comparisons and karyotypical studies. The aims of this study are not only to reconstruct the evolution of Brachypodium karyotypes to resolve the phylogeny, but also to highlight the mechanisms that shape the evolution of grass genomes. This was achieved through the use of comparative chromosome painting (CCP) which hybridises fluorescent, chromosome-specific probes derived from B. distachyon to homoeologous meiotic chromosomes of its close relatives. The study included five diploids (B. distachyon 2n = 10, B. sylvaticum 2n = 18, B. pinnatum 2n = 16; 2n = 18, B. arbuscula 2n = 18 and B. stacei 2n = 20) three allotetraploids (B. pinnatum 2n = 28, B. phoenicoides 2n = 28 and B. hybridum 2n = 30), and two species of unknown ploidy (B. retusum 2n = 38 and B. mexicanum 2n = 40). On the basis of the patterns of hybridisation and incorporating published data, we propose two alternative, but similar, models of karyotype evolution in the genus Brachypodium. According to the first model, the extant genome of B. distachyon derives from B. mexicanum or B. stacei by several rounds of descending dysploidy, and the other diploids evolve from B. distachyon via ascending dysploidy. The allotetraploids arise by interspecific hybridisation and chromosome doubling between B. distachyon and other diploids. The second model differs from the first insofar as it incorporates an intermediate 2n = 18 species between the B. mexicanum or B. stacei progenitors and the dysploidic B. distachyon

Ribosomal DNA, tri- and bi-partite pericentromeres in the permanent translocation heterozygote Rhoeo spathacea

High- and low-stringency FISH and base-specific fluorescence were performed on the permanent translocation heterozygote Rhoeo spathacea (2n = 12). Our results indicate that 45S rDNA arrays, rDNA-related sequences and other GC-rich DNA fraction(s) are located within the pericentromeric regions of all twelve chromosomes, usually colocalizing with the chromomycin A3-positive bands. Homogenization of the pericentromeric regions appears to result from the concerted spread of GC-rich sequences, with differential amplification likely. We found new 5S rDNA patterns, which suggest a variability in the breakpoints and in the consequent chromosome reorganizations. It was found that the large 5S rDNA locus residing on each of the 8E and 9E arms consisted of two smaller loci. On each of the two chromosome arms 3b and 4b, in addition to the major subtelomeric 5S rDNA locus, a new minor locus was found interstitially about 40% along the arm length. The arrangement of cytotogenetic landmarks and chromosome arm measurements are discussed with regard to genome repatterning in Rhoeo

Brachypodium distachyon - a useful model in the qualification of mutagen-induced micronuclei using multicolor FISH

Brachypodium distachyon (Brachypodium) is now intensively utilized as a model grass species in various biological studies. Its favorable cytological features create a unique foundation for a convenient system in mutagenesis, thereby potentially enabling the 'hot spots' and 'cold spots' of DNA damage in its genome to be analyzed. The aim of this study was to analyze the involvement of 5S rDNA, 25S rDNA, the Arabidopsis-type (TTTAGGG)n telomeric sequence and the Brachypodium-originated centromeric BAC clone CB33J12 in the micronuclei formation in Brachypodium root tip cells that were subjected to the chemical clastogenic agent maleic hydrazide (MH). To the best of our knowledge, this is the first use of a multicolor fluorescence in situ hybridization (mFISH) with four different DNA probes being used simultaneously to study plant mutagenesis. A quantitative analysis allowed ten types of micronuclei, which were characterized by the presence or absence of specific FISH signal(s), to be distinguished, thus enabling some specific rules governing the composition of the MH-induced micronuclei with the majority of them originating from the terminal regions of chromosomes, to be identified. The application of rDNA sequences as probes showed that 5S rDNA-bearing chromosomes are involved in micronuclei formation more frequently than the 25S rDNA-bearing chromosomes. These findings demonstrate the promising potential of Brachypodium to be a useful model organism to analyze the effects of various genotoxic agents on the plant nuclear genome stability, especially when the complex FISHbased and chromosome-specific approaches such as chromosome barcoding and chromosome painting will be applied in future studies

Cytomolecular analysis of ribosomal DNA evolution in a natural allotetraploid brachypodium hybridum and its putative ancestors - dissecting complex repetitive structure of intergenic spacers

Nucleolar dominance is an epigenetic phenomenon associated with nuclear 35S rRNA genes and consists in selective suppression of gene loci inherited from one of the progenitors in the allopolyploid. Our understanding of the exact mechanisms that determine this process is still fragmentary, especially in case of the grass species. This study aimed to shed some light on the molecular basis of this genome-specific inactivation of 35S rDNA loci in an allotetraploid Brachypodium hybridum (2n = 30), which arose from the interspecific hybridization between two diploid ancestors that were very similar to modern B. distachyon (2n = 10) and B. stacei (2n = 20). Using fluorescence in situ hybridization with 25S rDNA and chromosome-specific BAC clones as probes we revealed that the nucleolar dominance is present not only in meristematic root-tip cells but also in differentiated cell fraction of B. hybridum. Additionally, the intergenic spacers (IGSs) from both of the putative ancestors and the allotetraploid were sequenced and analyzed. The presumptive transcription initiation sites, spacer promoters and repeated elements were identified within the IGSs. Two different length variants, 2.3 and 3.5 kb, of IGSs were identified in B. distachyon and B. stacei, respectively, however only the IGS that had originated from B. distachyon-like ancestor was present in the allotetraploid. The amplification pattern of B. hybridum IGSs suggests that some genetic changes occurred in inactive B. stacei-like rDNA loci during the evolution of the allotetraploid. We hypothesize that their preferential silencing is an effect of structural changes in the sequence rather than just the result of the sole inactivation at the epigenetic leve

CD81 (Cluster of Differentiation 81)

Cluster of differentiation (CD81) is a type of protein, which is encoded by CD81 gene. Beside that CD81 is also known under other names such as Target of the Antiproliferative Antibody 1 (TAPA-1) and Tetraspanin-28 (TSPAN28). Location of CD81 is known to be on chromosome 11 (11p15.5), where it contains 15-20 bases in length. It is expressed mostly in cells of testis, ovary, endometrium, placenta, bone marrow, smooth muscles and others. The main function of the CD81 protein is to mediate signal transduction events, which are important for cells' development, activation, growth and motility. The CD81 gene is also known as a candidate for many malignancies because of its location. The characteristic feature of CD81 is that it is highly hydrophobic and contains a short N- and C-terminal cytoplasmic domains together with cytoplasmic cysteines, potential sites of palmitoylation as well as four transmembrane domains where they together hold the protein in a cell membrane. There are two CD81 isoforms, isoform 1 and isoform 2. Isoforms of CD81 are usually found in a tumor-suppressor region where they have a great impact on tumor development. There has always been a high interest in research on CD81 function in viral disease development. In fact, it is known that CD81 contributes in the development of diseases such as hepatitis C, malaria and various types of cancer. Since the complete effect of CD81 is unknown, further research and scientific methodology could potentially discover all possible functions and mechanisms regulated by the CD81 protein in human body

Spatial distribution of centromeres and telomeres at interphase varies among Brachypodium species

In this study the 3-D distribution of centromeres and telomeres was analysed in the interphase nuclei of three Brachypodium species, i.e. B. distachyon (2n=10), B. stacei (2n=20) and B. hybridum (2n=30), which is presumably a hybrid between the first two species. Using fluorescence in situ hybridization (FISH) with centromeric and telomeric DNA probes, it was observed that the majority of B. distachyon nuclei in the root tip cells displayed the Rabl configuration while both B. stacei and B. hybridum mostly lacked the centromere-telomere polarization. In addition, differentiated leaf cells of B. distachyon did not display the Rabl pattern. In order to analyse the possible connection between the occurrence of the Rabl pattern and the phase of cell cycle or DNA content, FISH was combined with digital image cytometry. The results revealed that the frequency of nuclei with the Rabl configuration in the root tip nuclei was positively correlated with an increase in DNA content, which resulted from DNA replication. Also, the analysis of the influence of the nuclear shape on the nuclear architecture indicated that an increasing elongation of the nuclei negatively affected the occurrence of the Rabl pattern. Some possible explanations of these phenomena are discussed

Gas Purity Analytics, Calibration Studies, and Background Predictions towards the First Results of XENON1T

The XENON1T experiment aims at the direct detection of the well motivated dark matter candidate of weakly interacting massive particles (WIMPs) scattering off xenon nuclei. The first science run of 34.2 live days has already achieved the most stringent upper limit on spin-independent WIMP-nucleon cross-sections above masses of 10GeV with a minimum of 7.7x10^(-47) cm² at a mass of 35GeV. Crucial for this unprecedented sensitivity are a high xenon gas purity and a good understanding of the background. In this work, a procedure is described that was developed to measure the purity of the experiment’s xenon inventory of more than three tons during its initial transfer to the detector gas system. The technique of gas chromatography has been employed to analyze the noble gas for impurities with the focus on oxygen and krypton contaminations. Furthermore, studies on the calibration of the experiment’s dominating background induced by natural gamma and beta radiation were performed. Hereby, the novel sources of radioactive isotopes that can be dissolved in the xenon were employed, namely 220Rn and tritium. The sources were analyzed in terms of a potential impact on the outcome of a dark matter search. As a result of the promising findings for 220Rn, the source was successfully deployed in the first science run of XENON1T. The first WIMP search of XENON1T is outlined in this thesis, in which a background component from interactions taking place in close proximity to the detector wall is identified, investigated and modeled. A background prediction was derived that was incorporated into the background model of the WIMP search which was found to be in good agreement with the observation