The role of RNA primer in discontinuous DNA replication in isolated nuclei from HeLa cells

RNA-primed discontinuous DNA synthesis was studied in an in vitro system consisting of washed nuclei from synchronized S-phase HeLa cells. A new technique proved useful for the purification of short nascent fragments of DNA (Okazaki fragments). Mercurated dCTP was substituted for dCTP in the DNA synthesis reaction. Short nascent pieces (4–6 S) of mercurated DNA were found to bind preferentially to sulfhydryl-agarose, and could be eluted with mercaptoethanol. The isolated fragments were assayed for the presence of covalently linked RNA by the spleen exonuclease method described by Kurosawa et al. (Kurosawa, Y., Ogawa, T., Hirose, S., Okazaki, T. and Okazaki, R. (1975) J. Mol. Biol. 96, 653–664). Following a 30 s incubation with [³H]TTP in the absence of added ribonucleotides, approximately 20% of the nascent strands synthesized in washed nuclear preparations had RNA attached. These RNA primers either preexisted in the nuclei or were formed from endogenous ribonucleotides. The 5′ ends of the primers appeared to be largely in a phosphorylated state. In the absence of added ribonucleotides, these RNA-DNA linkages disappeared within 2 min, whereas if ribonucleotides were added, the number of RNA primers increased to 40% and remained at this level for greater than 2 min. To obtain maximal levels of RNA primer, the addition of all three of the ribonucleotides, rCTP, rGTP and rUTP (0.1 mM), as well as high levels of rATP (5 mM) was required. Addition of ribonucleotides also markedly enhanced the amount of nascent DNA fragments synthesized. However, in the absence of added ribonucleotides, after RNA primers had disappeared, nascent DNA fragments were still initiated at a significant rate. These results suggest that RNA primers play an important role in the initiation of Okazaki fragments but that synthesis can also be initiated by alternative mechanisms. An important role for ATP in RNA primer synthesis is suggested.     

Evaluation of genotoxicity of combined soil pollution by cadmium and imidacloprid

Cadmium (Cd) is one of the important pollutants of soil and the genotoxicity of Cd-contaminated soil was studied in combination with imidacloprid. The single cell gel electrophoresis or comet assay was used to quantify DNA strand breaks as a measure of DNA damage induced by Cd and imidacloprid contamination in soil. The soil was artificially contaminated by Cd 2 h at 25℃ and were used in the comet assay. DNA damage was measured as the values of percentage of nuclei with tails, tail length, tail DNA, tail moment (TM), and Olive tail moment (OTM). DNA damages of root tips of Vicia faba increased after Cd treatment and there were dose-related increases in DNA damage measured as these parameters. However, the addition of imidacloprid further increased the DNA damage. These data confirmed the genotoxic effect of Cd to plants, and that the combined pollution with imidacloprid can enhance the genotoxicity of Cd.     

Odourant dominance in olfactory mixture processing: what makes a strong odourant?

The question of how animals process stimulus mixtures remains controversial as opposing views propose that mixtures are processed analytically, as the sum of their elements, or holistically, as unique entities different from their elements. Overshadowing is a widespread phenomenon that can help decide between these alternatives. In overshadowing, an individual trained with a binary mixture learns one element better at the expense of the other. Although element salience (learning success) has been suggested as a main explanation for overshadowing, the mechanisms underlying this phenomenon remain unclear. We studied olfactory overshadowing in honeybees to uncover the mechanisms underlying olfactory-mixture processing. We provide, to our knowledge, the most comprehensive dataset on overshadowing to date based on 90 experimental groups involving more than 2700 bees trained either with six odourants or with their resulting 15 binary mixtures. We found that bees process olfactory mixtures analytically and that salience alone cannot predict overshadowing. After normalizing learning success, we found that an unexpected feature, the generalization profile of an odourant, was determinant for overshadowing. Odourants that induced less generalization enhanced their distinctiveness and became dominant in the mixture. Our study thus uncovers features that determine odourant dominance within olfactory mixtures and allows the referring of this phenomenon to differences in neural activity both at the receptor and the central level in the insect nervous system.     

Species introduction through seeds from an old,species‐rich hay meadow: Effects of management

Abstract. A local seed mixture from plants growing in a species‐rich, traditionally managed hay meadow site at Varaldsoy, Hardanger, western Norway, where many endangered hay meadow species of the region are growing, was sown in a newly harrowed experimental field 1 km from the source meadow in order to increase the habitat area for the endangered species. Of 25 endangered species recorded in the source meadow, only one (Holcus lanatus) was present in the target meadow. After sowing, 16 of the endangered species in addition to Holcus lanatus were recorded in the new site. Six species were only present in sown plots and seven others were more frequent there, while three species might have arrived by chance or originated from the seed bank. Replacing the traditional management regime, including one late cut and grazing in spring and in autumn, with three cutting times and the creation of gaps in the sward, resulted in a higher number of endangered species in plots which were only cut, possibly because the grazing was too intensive in the small enclosures.     

Blood-based profiles of DNA methylation predict the underlying distribution of cell types

The potential influence of underlying differences in relative leukocyte distributions in studies involving blood-based profiling of DNA methylation is well recognized and has prompted development of a set of statistical methods for inferring changes in the distribution of white blood cells using DNA methylation signatures. However, the extent to which this methodology can accurately predict cell-type proportions based on blood-derived DNA methylation data in a large-scale epigenome-wide association study (EWAS) has yet to be examined. We used publicly available data deposited in the Gene Expression Omnibus (GEO) database (accession number GSE37008), which consisted of both blood-derived epigenome-wide DNA methylation data assayed using the Illumina Infinium HumanMethylation27 BeadArray and complete blood cell (CBC) counts among a community cohort of 94 non-diseased individuals. Constrained projection (CP) was used to obtain predictions of the proportions of lymphocytes, monocytes and granulocytes for each of the study samples based on their DNA methylation signatures. Our findings demonstrated high consistency between the average CBC-derived and predicted percentage of monocytes and lymphocytes (17.9% and 17.6% for monocytes and 82.1% and 81.4% for lymphocytes), with root mean squared error (rMSE) of 5% and 6%, for monocytes and lymphocytes, respectively. Similarly, there was moderate-high correlation between the CP-predicted and CBC-derived percentages of monocytes and lymphocytes (0.60 and 0.61, respectively), and these results were robust to the number of leukocyte differentially methylated regions (L-DMRs) used for CP prediction. These results serve as further validation of the CP approach and highlight the promise of this technique for EWAS where DNA methylation is profiled using whole-blood genomic DNA.     

Migration stopover ecology of Cinnamon Teal in western North America

Identifying migration routes and fall stopover sites of Cinnamon Teal (Spatula cyanoptera septentrionalium) can provide a spatial guide to management and conservation efforts, and address vulnerabilities in wetland networks that support migratory waterbirds. Using high spatiotemporal resolution GPS‐GSM transmitters, we analyzed 61 fall migration tracks across western North America during our three‐year study (2017–2019). We marked Cinnamon Teal primarily during spring/summer in important breeding and molting regions across seven states (California, Oregon, Washington, Idaho, Utah, Colorado, and Nevada). We assessed fall migration routes and timing, detected 186 fall stopover sites, and identified specific North American ecoregions where sites were located. We classified underlying land cover for each stopover site and measured habitat selection for 12 land cover types within each ecoregion. Cinnamon Teal selected a variety of flooded habitats including natural, riparian, tidal, and managed wetlands; wet agriculture (including irrigation ditches, flooded fields, and stock ponds); wastewater sites; and golf and urban ponds. Wet agriculture was the most used habitat type (29.8% of stopover locations), and over 72% of stopover locations were on private land. Relatively scarce habitats such as wastewater ponds, tidal marsh, and golf and urban ponds were highly selected in specific ecoregions. In contrast, dry non‐habitat across all ecoregions, and dry agriculture in the Cold Deserts and Mediterranean California ecoregions, was consistently avoided. Resources used by Cinnamon Teal often reflected wetland availability across the west and emphasize their adaptability to dynamic resource conditions in arid landscapes. Our results provide much needed information on spatial and temporal resource use by Cinnamon Teal during migration and indicate important wetland habitats for migrating waterfowl in the western United States.     

Calculation of Vapour-Liquid Equilibria of Lennard-Jones Binary Systems by the Gibbs Ensemble Monte Carlo Simulation

Abstract

The Gibbs ensemble Monte Carlo simulation has been used to calculate vapour-liquid equilibria of a Lennard-Jones (LJ) binary mixture. The mixture studied is the LB-2-1 model which has been used in our previous calculations on PVT relation and density-dependent local composition. The P-x-y relation has been established at two different temperatures and used to determine vapour-liquid coexistence region in the PVTx space.     

Mixture design of starchy substrates hydrolysis by an immobilized glucoamylase from Aspergillus brasiliensis

Starch has great importance in human diet, since it is a heteropolymer of plants, mainly found in roots, as potato, cassava and arrowroots. This carbohydrate is composed by a highly-branched chain: amylopectin; and a linear chain: amylose. The proportion between the chains varies according to the botanical source. Starch hydrolysis is catalyzed by enzymes of the amilolytic system, named amylases. Among the various enzymes of this system, the glucoamylases (EC 3.2.1.3 glucan 1,4-alpha-glucosidases) are the majority because they hydrolyze the glycosidic linkages at the end of starch chains releasing glucose monomers. In this work, a glucoamylase secreted in the culture medium, by the ascomycete Aspergillus brasiliensis, was immobilized in Dietilaminoetil Sepharose-Polyethylene Glycol (DEAE-PEG), since immobilized biocatalysts are more stable in long periods of hydrolysis, and can be recovered from the final product and reused for several cycles. Glucoamylase immobilization has shown great thermal stability improvement over the soluble enzyme, reaching 66% more activity after 6?h at 60?°C, and 68% of the activity after 10 hydrolysis cycles. A simplex centroid experimental mixture design was applied as a tool to characterize the affinity of the immobilized enzyme for different starchy substrates. In assays containing several proportions of amylose, amylopectin and starch, the glucoamylase from A. brasiliensis mainly hydrolyzed the amylopectin chains, showing to have preference by branched substrates.     

Spectral feature differences between shrub and grass communities and shrub coverage retrieval in shrub-encroached grassland in Xianghuang Banner,Nei Mongol,China北大核心CSCD

Aims: Shrub-encroached grassland has become an important vegetation type in China's arid and semi-arid region. Our study objective is to explore the spectral features of shrub and grass communities, as well as their empirical relationships with shrub coverage. The quantitative estimation of shrub cover based on medium-resolution Landsat satellite imagery provides the practical basis for long term retrieval of large areas of shrub expansion in the grassland region. Methods: Linear models and Multiple Endmember Spectral Analysis Model (MESMA) based on medium resolution Landsat satellite imagery were developed to quantify the shrub coverage in a shrub-encroached grassland region in Xianghuang Banner, Nei Mongol using the spectral features and their seasonal differences between the shrub and grass communities. Important findings: Compared to Leymus chinensis and Stipa krylovii dominated grass communities, Caragana microphylla community had a higher normalized difference vegetation index (NDVI), modified red edge normalized difference vegetation index (mNDVI705), and red edge slope. The red edge position of C. microphylla community shifted to longer wavelengths. The average and the maximum shrub coverage was 13% and 25%, respectively, in the shrub-encroached grassland based on both models. The correlation coefficient of determination (R2) and root mean square error (RMSE) of the linear model was 0.31 and 0.05, respectively. We found that the linear model based on seasonal differences of shrub and grass community was more suitable for retrieving shrub coverage in the study area from medium resolution imagery than the MESMA model that is based on mid-summer images.