Genetic diversity among varieties and wild species accessions of pea (Pisum sativum L.) based on SSR markers

To assess the genetic relations inPisum genus and to examine putative duplicate accessions, 20 pea varieties (Pisum sativum L.) with 57 accessions from wild Pisum species fulvum, subspecies (subsp.) asiaticum, elatius, thebaicum, abyssinicum, transcaucasicum and arvense were analyzed using 10 out of 20 microsatellite primer pairs. We genotyped all accessions. In total, 59 alleles were identified in whole collection. The maximum number of alleles (8 alleles) was obtained from the PEACPLHPP, AF004843, and AA43090 loci. The maximum number of private alleles (4) in the wild collection was detected in AF004843 locus but in the cultivar collection, it was detected in AA430902 and PSBLOX13.2 loci. Cluster analysis and principal coordinate analysis located accessions in 3 groups and cultivated varieties were obviously separated from the wild accessions. Analysis of molecular variance (AMOVA) revealed that the intergroups component of variance (29%) is lower than the intragroups component of variance (71%). The lowest value of genetic differentiation ( Pisum genus and to examine putative duplicate accessions, 20 pea varieties (Pisum sativum L.) with 57 accessions from wild Pisum species fulvum, subspecies (subsp.) asiaticum, elatius, thebaicum, abyssinicum, transcaucasicum and arvense were analyzed using 10 out of 20 microsatellite primer pairs. We genotyped all accessions. In total, 59 alleles were identified in whole collection. The maximum number of alleles (8 alleles) was obtained from the PEACPLHPP, AF004843, and AA43090 loci. The maximum number of private alleles (4) in the wild collection was detected in AF004843 locus but in the cultivar collection, it was detected in AA430902 and PSBLOX13.2 loci. Cluster analysis and principal coordinate analysis located accessions in 3 groups and cultivated varieties were obviously separated from the wild accessions. Analysis of molecular variance (AMOVA) revealed that the intergroups component of variance (29%) is lower than the intragroups component of variance (71%). The lowest value of genetic differentiation (Pisum genus and to examine putative duplicate accessions, 20 pea varieties (Pisum sativum L.) with 57 accessions from wild Pisum species fulvum, subspecies (subsp.) asiaticum, elatius, thebaicum, abyssinicum, transcaucasicum and arvense were analyzed using 10 out of 20 microsatellite primer pairs. We genotyped all accessions. In total, 59 alleles were identified in whole collection. The maximum number of alleles (8 alleles) was obtained from the PEACPLHPP, AF004843, and AA43090 loci. The maximum number of private alleles (4) in the wild collection was detected in AF004843 locus but in the cultivar collection, it was detected in AA430902 and PSBLOX13.2 loci. Cluster analysis and principal coordinate analysis located accessions in 3 groups and cultivated varieties were obviously separated from the wild accessions. Analysis of molecular variance (AMOVA) revealed that the intergroups component of variance (29%) is lower than the intragroups component of variance (71%). The lowest value of genetic differentiation (PT = 0.27) of pair wise collections between wild and variety collections, was detected in ssp. elatius. Assignment test on the basis of log-likelihood to estimate the likelihood that an individual belongs to a given group, showed that 96% of accessions being assigned correctly to their groups. This study showed that genetic probability profiles of accessions can corroborate clustering analyses while providing additional information as a powerful tool for assigning accessions into their related groups

Validation of a genus-specific gene; TPS, used as internal control in quantitative Real Time PCR of transgenic cotton

Abstract Identification of genes with invariant levels of gene expression is a prerequisite for validating transcriptomic changes accompanying development. Ideally expression of these genes should be independent of the morphogenetic process or environmental condition.We report here the validation of internal control gene i.e.TPS (trehalose 6-phosphate-synthase) in cotton (Gossypium spp), using TaqMan system in quantitative Real Time PCR (qRT-PCR). The Gene expression was tested in five different G. hirsutum cultivars including Coker 312, Acala SJ, ZETA 2, Taghva, Neishabour and a diploid wild type; G. barbadense. Identical amplicons were obtained within these cultivars. No amplifications was achieved when DNA samples from barley (Hordeum vulgare), maize (Zea mays), rice (Oryza sativa)

The extent of error-prone replication-restart by homologous recombination is controlled by Exo1 and checkpoint proteins

Genetic instability, a hallmark of cancer, can occur when the replication machinery encounters a barrier. The intra-S phase checkpoint maintains stalled replication forks in a replication-competent configuration by phosphorylating replisome components and DNA repair proteins to prevent forks from catastrophically collapsing. Here we report a novel Chk1- and Cds1Chk2-independent function for Rad3ATR, the core S. pombe checkpoint sensor kinase: Rad3ATR regulates the association of recombination factors with collapsed forks thus limiting their genetic instability. We further reveal antagonistic roles for Rad3ATR and the 9-1-1 clamp: Rad3ATR restrains MRN- and Exo1-dependent resection while the 9-1-1 complex promotes Exo1 activity. Interestingly the MRN complex, but not its nuclease activity, promotes resection and the subsequent association of recombination factors at collapsed forks. The biological significance of this regulation is revealed by the observation that Rad3ATR prevents Exo1-dependent genome instability upstream a collapsed fork without affecting the efficiency of recombination-mediated replication-restart. We propose the interplay between Rad3ATR and the 9-1-1 clamp functions to fine-tune the balance between the need for recovery of replication via recombination and the risk of increased genome instability

A truncated DNA-damage-signaling response is activated after DSB formation in the G1 phase of Saccharomyces cerevisiae

In Saccharomyces cerevisiae, the DNA damage response (DDR) is activated by the spatio-temporal colocalization of Mec1-Ddc2 kinase and the 9-1-1 clamp. In the absence of direct means to monitor Mec1 kinase activation in vivo, activation of the checkpoint kinase Rad53 has been taken as a proxy for DDR activation. Here, we identify serine 378 of the Rad55 recombination protein as a direct target site of Mec1. Rad55-S378 phosphorylation leads to an electrophoretic mobility shift of the protein and acts as a sentinel for Mec1 activation in vivo. A single double-stranded break (DSB) in G1-arrested cells causes phosphorylation of Rad55-S378, indicating activation of Mec1 kinase. However, Rad53 kinase is not detectably activated under these conditions. This response required Mec1-Ddc2 and loading of the 9-1-1 clamp by Rad24-RFC, but not Rad9 or Mrc1. In addition to Rad55–S378, two additional direct Mec1 kinase targets are phosphorylated, the middle subunit of the ssDNA-binding protein RPA, RPA2 and histone H2A (H2AX). These data suggest the existence of a truncated signaling pathway in response to a single DSB in G1-arrested cells that activates Mec1 without eliciting a full DDR involving the entire signaling pathway including the effector kinases

Mechanisms of carbon dioxide detection in the earthworm Dendrobaena veneta

IntroductionCarbon dioxide (CO2) is a critical biological signal that is noxious to many animals at high concentrations. The earthworm Dendrobaena veneta lives in subterranean burrows containing high levels of CO2 and respires through its skin. Despite the ecological and agricultural importance of earthworms, relatively little is known about how they make decisions in their environment, including their response to elevated levels of CO2.MethodsTo examine CO2 detection in this species, we designed the exudate assay, in which we placed an earthworm in a sealed container, exposed it to varying concentrations of CO2 for one minute, and recorded the amount of exudate secreted. Because earthworms excrete exudate in response to noxious stimuli, we hypothesized that the amount of exudate produced was proportional to the amount of irritation. We repeated these experiments after treatment with several blockers for molecules with potential involvement in CO2 detection, including carbonic anhydrases, guanylate cyclase, TRPA1, ASICs, and OTOP channels. We also confirmed the presence of homologous transcripts for each of these gene families in an epithelial transcriptome for D. veneta. Additionally, since organisms often detect CO2 levels indirectly by monitoring the conversion to carbonic acid (a weak acid), we used the exudate assay to evaluate aversion to additional weak acids (formic acid, acetic acid, and propionic acid).ResultsEarthworms excreted significantly more exudate in response to CO2 in a dosage-dependent manner, and this response was muted by the general carbonic anhydrase inhibitor acetazolamide, the carbonic anhydrase IX/XII inhibitor indisulam, the calcium channel blocker ruthenium red, the sodium channel blocker amiloride, and the acid-sensing ion channel blocker diminazene aceturate.DiscussionThese data provide evidence of the role of carbonic anhydrase and epithelial sodium channels in earthworm CO2 detection, establish that, similar to other subterranean-dwelling animals, earthworms are extremely tolerant of CO2, and contribute to our understanding of the mechanisms used by earthworms to detect and react to weak acids in their environment

The DNA damage checkpoint pathways exert multiple controls on the efficiency and outcome of the repair of a double-stranded DNA gap

A DNA gap repair assay was used to determine the effect of mutations in the DNA damage checkpoint system on the efficiency and outcome (crossover/non-crossover) of recombinational DNA repair. In Saccharomyces cerevisiae gap repair is largely achieved by homologous recombination. As a result the plasmid either integrates into the chromosome (indicative of a crossover outcome) or remains extrachromosomal (indicative of a non-crossover outcome). Deletion mutants of the MEC1 and RAD53 checkpoint kinase genes exhibited a 5-fold decrease in gap repair efficiency, showing that 80% of the gap repair events depended on functional DNA damage checkpoints. Epistasis analysis suggests that the DNA damage checkpoints affect gap repair by modulating Rad51 protein-mediated homologous recombination. While in wild-type cells only ∼25% of the gap repair events were associated with a crossover outcome, Mec1-deficient cells exhibited a >80% crossover association. Also mutations in the effector kinases Rad53, Chk1 and Dun1 were found to affect crossover association of DNA gap repair to various degrees. The data suggest that the DNA damage checkpoints are important for the optimal functioning of recombinational DNA repair with multiple terminal targets to modulate the efficiency and outcome of homologous recombination

Design of LVDC bidirectional hybrid circuit breaker

One of the most challenging hindrances to the spread use of the DC grids is their protection devices. Due to the absence of zero crossing, when a fault occurs, arc extinction becomes an issue for the traditional mechanical circuit breakers. Thanks to the recent advances in solid-state circuit breakers, this problem is solved because of the absence of mechanical contacts in their topology. However, these breakers cause not negligible conduction losses. This paper illustrates a new low-voltage bidirectional hybrid circuit breaker able to interrupt short-circuit current in few milliseconds. It provides a low-loss path for the current during the normal conduction and a fast current interruption with a limited arc level in case of fault occurrence. The working principle of the device is explained, and its functionality is proven through simulation results

Dynamic assessment of the impacts of global warming on nitrate losses from a subsurface-drained rainfed-canola field

The impact of global warming on water and nitrate losses from a rainfed-canola cropping system under various artificial drainage systems was assessed using an integrated field-modeling approach. Four subsurface drainage systems with different drain depths (Dx) and spacings (Ly), including D0.90L30, D0.65L30, D0.65L15, and Bilevel (with a drain spacing of 15 m and alternate drain depths of 0.65 and 0.90 m), were considered. The HYDRUS (2D/3D) model was first calibrated and validated using data collected for all drainage systems during the 2015–2016 and 2016–2017 canola cropping cycles, respectively, and then applied to simulate water/nitrate losses for different drainage systems under meteorological conditions predicted assuming expected future global warming. Future weather data were downscaled from 20 general circulation models and four RCP scenarios for the mid 21st century (for 2041–2070). The model capability of representing experimental field data was evaluated using the mean bias error (MBE), the normalized root mean square error (nRMSE), and the model efficiency (EF). The HYDRUS (2D/3D) model provided reliable description of soil water contents (MBE=-0.5 % to 0.2 %, nRMSE = 0.005−0.034%, and EF = 0.73−0.99), drainage fluxes (MBE= -21.7 × 10−3 to 24.9 × 10−3 mm d-1, nRMSE = 0.23−0.37%, and EF = 0.69−0.85), soil nitrate concentrations (MBE= -0.002 to 1.00 mg cm−3, nRMSE = 0.08−0.18%, and EF = 0.51−0.88), and nitrate fluxes (MBE= -0.97 to 0.72 mg cm-1 d-1, nRMSE = 0.35−0.57%, and EF = 0.77−0.87). The modeling results indicate that climate change will cause an increase of up to 148 % in average daily drainage fluxes and up to 125 % in average daily nitrate fluxes compared to the base case. This will result in an increase of 4–125 % in seasonal nitrate losses from various drainage systems, with the lowest and highest projections for the D0.65L15 and D0.65L30 systems, respectively. The HYDRUS-simulated results indicate that the D0.65L15 system is environmentally safer than the other evaluated drainage systems for predicted global warming conditions concerning water/nitrate losses