Counting bubbles : precision process control of gas-liquid reactions in flow with an optical inline sensor

Quality by Design encouraged by the US Food and Drug Administration (FDA) in the continuous flow synthesis requires tight monitoring of all the reaction input and output parameters to improve reproducibility and eliminate the process rejects. The reaction monitoring, however, relies on costly (above 10,000$) process analytical technology (PAT) – one of the factors that prevents a wider utilisation of continuous processes. In the work, we show that gas-liquid reactions can be monitored using low-cost (10$) hardware – optical liquid inline sensors – that allows instantaneous analysis of gas fraction in the moving stream. We discuss the application of the sensor for various gas-liquid reactions. The gas-consuming reactions such as hydrogenation are the easiest to implement because the sensor without calibration provides accurate readings close to complete consumption of the gas. The gas-evolving reactions can be monitored but require sensor calibration to determine the gas fraction accurately. Operation of the sensor was demonstrated for various hydrogenation reactions self-optimised using a proportional-integral (PID) algorithm which adjusted the substrate concentration to provide high (but not full) pre-defined hydrogen consumption. The optimised hydrogen consumption agreed with the product analysis for a range of the substrates hydrogenated under various pressures and with different selectivities. The optical sensor was also proven to be an efficient tool in adapting the reaction condition to the catalyst deactivation in the reaction of 2-methyl-3-butyn-2-ol semi-hydrogenation – the autonomous reactor allowed reaching a turn-over number (TON) of 2.7·106 with the value of 1.5·107 expected till a twofold decrease in the catalyst activity. The TON values demonstrated are significantly higher than those observed in batch reactors (~103) even in case of catalyst re-use (105) demonstrating a substantial improvement of process sustainability operating with the process control

Power system dynamic state estimation: motivations, definitions, methodologies, and future work

This paper summarizes the technical activities of the Task Force on Power System Dynamic State and Parameter Estimation. This Task Force was established by the IEEE Working Group on State Estimation Algorithms to investigate the added benefits of dynamic state and parameter estimation for the enhancement of the reliability, security, and resilience of electric power systems. The motivations and engineering values of dynamic state estimation (DSE) are discussed in detail. Then, a set of potential applications that will rely on DSE is presented and discussed. Furthermore, a unified framework is proposed to clarify the important concepts related to DSE, forecasting-aided state estimation, tracking state estimation, and static state estimation. An overview of the current progress in DSE and dynamic parameter estimation is provided. The paper also provides future research needs and directions for the power engineering community

Analysis of the P. lividus sea urchin genome highlights contrasting trends of genomic and regulatory evolution in deuterostomes

Sea urchins are emblematic models in developmental biology and display several characteristics that set them apart from other deuterostomes. To uncover the genomic cues that may underlie these specificities, we generated a chromosome-scale genome assembly for the sea urchin Paracentrotus lividus and an extensive gene expression and epigenetic profiles of its embryonic development. We found that, unlike vertebrates, sea urchins retained ancestral chromosomal linkages but underwent very fast intrachromosomal gene order mixing. We identified a burst of gene duplication in the echinoid lineage and showed that some of these expanded genes have been recruited in novel structures (water vascular system, Aristotle's lantern, and skeletogenic micromere lineage). Finally, we identified gene-regulatory modules conserved between sea urchins and chordates. Our results suggest that gene-regulatory networks controlling development can be conserved despite extensive gene order rearrangement

Selection and Validation of Reference Genes for Quantitative Real-Time PCR in Buckwheat (Fagopyrum esculentum) Based on Transcriptome Sequence Data

Quantitative reverse transcription PCR (qRT-PCR) is one of the most precise and widely used methods of gene expression analysis. A necessary prerequisite of exact and reliable data is the accurate choice of reference genes. We studied the expression stability of potential reference genes in common buckwheat (Fagopyrum esculentum) in order to find the optimal reference for gene expression analysis in this economically important crop. Recently sequenced buckwheat floral transcriptome was used as source of sequence information. Expression stability of eight candidate reference genes was assessed in different plant structures (leaves and inflorescences at two stages of development and fruits). These genes are the orthologs of Arabidopsis genes identified as stable in a genome-wide survey gene of expression stability and a traditionally used housekeeping gene GAPDH. Three software applications – geNorm, NormFinder and BestKeeper - were used to estimate expression stability and provided congruent results. The orthologs of AT4G33380 (expressed protein of unknown function, Expressed1), AT2G28390 (SAND family protein, SAND) and AT5G46630 (clathrin adapter complex subunit family protein, CACS) are revealed as the most stable. We recommend using the combination of Expressed1, SAND and CACS for the normalization of gene expression data in studies on buckwheat using qRT-PCR. These genes are listed among five the most stably expressed in Arabidopsis that emphasizes utility of the studies on model plants as a framework for other species

Identification of superior reference genes for data normalisation of expression studies via quantitative PCR in hybrid roses (Rosa hybrida)

<p>Abstract</p> <p>Background</p> <p>Gene expression studies are a prerequisite for understanding the biological function of genes. Because of its high sensitivity and easy use, quantitative PCR (qPCR) has become the gold standard for gene expression quantification. To normalise qPCR measurements between samples, the most prominent technique is the use of stably expressed endogenous control genes, the so called reference genes. However, recent studies show there is no universal reference gene for all biological questions. Roses are important ornamental plants for which there has been no evaluation of useful reference genes for gene expression studies.</p> <p>Results</p> <p>We used three different algorithms (BestKeeper, geNorm and NormFinder) to validate the expression stability of nine candidate reference genes in different rose tissues from three different genotypes of <it>Rosa hybrida </it>and in leaves treated with various stress factors. The candidate genes comprised the classical "housekeeping genes" (<it>Actin, EF-1α, GAPDH</it>, <it>Tubulin </it>and <it>Ubiquitin</it>), and genes showing stable expression in studies in <it>Arabidopsis </it>(<it>PP2A, SAND, TIP </it>and <it>UBC</it>). The programs identified no single gene that showed stable expression under all of the conditions tested, and the individual rankings of the genes differed between the algorithms. Nevertheless the new candidate genes, specifically, <it>PP2A </it>and <it>UBC</it>, were ranked higher as compared to the other traditional reference genes. In general, <it>Tubulin </it>showed the most variable expression and should be avoided as a reference gene.</p> <p>Conclusions</p> <p>Reference genes evaluated as suitable in experiments with <it>Arabidopsis thaliana </it>were stably expressed in roses under various experimental conditions. In most cases, these genes outperformed conventional reference genes, such as <it>EF1-α </it>and <it>Tubulin</it>. We identified <it>PP2A</it>, <it>SAND </it>and <it>UBC </it>as suitable reference genes, which in different combinations may be used for normalisation in expression analyses via qPCR for different rose tissues and stress treatments. However, the vast genetic variation found within the genus <it>Rosa</it>, including differences in ploidy levels, might also influence expression stability of reference genes, so that future research should also consider different genotypes and ploidy levels.</p

Pathogen-induced activation of disease-suppressive functions in the endophytic root microbiome

Microorganisms living inside plants can promote plant growth and health, but their genomic and functional diversity remain largely elusive. Here, metagenomics and network inference show that fungal infection of plant roots enriched for Chitinophagaceae and Flavobacteriaceae in the root endosphere and for chitinase genes and various unknown biosynthetic gene clusters encoding the production of nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs). After strain-level genome reconstruction, a consortium of Chitinophaga and Flavobacterium was designed that consistently suppressed fungal root disease. Site-directed mutagenesis then revealed that a previously unidentified NRPS-PKS gene cluster from Flavobacterium was essential for disease suppression by the endophytic consortium. Our results highlight that endophytic root microbiomes harbor a wealth of as yet unknown functional traits that, in concert, can protect the plant inside out.Microbial Biotechnolog

Circulating tumor cells criteria (CyCAR) versus standard RECIST criteria for treatment response assessment in metastatic colorectal cancer patients

The use of circulating tumor cells (CTCs) as indicators of treatment response in metastatic colorectal cancer (mCRC) needs to be clarified. The objective of this study is to compare the Response Evaluation Criteria in Solid Tumors (RECIST) with the Cytologic Criteria Assessing Response (CyCAR), based on the presence and phenotypic characterization of CTCs, as indicators of FOLFOX–bevacizumab treatment response. We observed a decrease of CTCs (42.8 vs. 18.2%) and VEGFR positivity (69.7% vs. 41.7%) after treatment. According to RECIST, 6.45% of the patients did not show any clinical benefit, whereas 93.55% patients showed a favorable response at 12 weeks. According to CyCAR, 29% had a non-favorable response and 71% patients did not. No significant differences were found between the response assessment by RECIST and CyCAR at 12 or 24 weeks. However, in the multivariate analysis, RECIST at 12 weeks and CyCAR at 24 weeks were independent prognostic factors for OS (HR: 0.1, 95% CI 0.02–0.58 and HR: 0.35, 95% CI 0.12–0.99 respectively). CyCAR results were comparable to RECIST in evaluating the response in mCRC and can be used as an alternative when the limitation of RECIST requires additional response analysis techniques.This work was supported by Roche Spain and a Ph.D. grant from the University of Granada

Reference gene validation for quantitative RT-PCR during biotic and abiotic stresses in Vitis vinifera

Grapevine is one of the most cultivated fruit crop worldwide with Vitis vinifera being the species with the highest economical importance. Being highly susceptible to fungal pathogens and increasingly affected by environmental factors, it has become an important agricultural research area, where gene expression analysis plays a fundamental role. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is currently amongst the most powerful techniques to perform gene expression studies. Nevertheless, accurate gene expression quantification strongly relies on appropriate reference gene selection for sample normalization. Concerning V. vinifera, limited information still exists as for which genes are the most suitable to be used as reference under particular experimental conditions. In this work, seven candidate genes were investigated for their stability in grapevine samples referring to four distinct stresses (Erysiphe necator, wounding and UV-C irradiation in leaves and Phaeomoniella chlamydospora colonization in wood). The expression stability was evaluated using geNorm, NormFinder and BestKeeper. In all cases, full agreement was not observed for the three methods. To provide comprehensive rankings integrating the three different programs, for each treatment, a consensus ranking was created using a non-weighted unsupervised rank aggregation method. According to the last, the three most suitable reference genes to be used in grapevine leaves, regardless of the stress, are UBC, VAG and PEP. For the P. chlamydospora treatment, EF1, CYP and UBC were the best scoring genes. Acquaintance of the most suitable reference genes to be used in grapevine samples can contribute for accurate gene expression quantification in forthcoming studiesinfo:eu-repo/semantics/publishedVersio

Spindle Assembly Checkpoint Protein Dynamics Reveal Conserved and Unsuspected Roles in Plant Cell Division

Background: In eukaryotes, the spindle assembly checkpoint (SAC) ensures that chromosomes undergoing mitosis do not segregate until they are properly attached to the microtubules of the spindle. Methodology/Principal Findings: We investigated the mechanism underlying this surveillance mechanism in plants, by characterising the orthogolous SAC proteins BUBR1, BUB3 and MAD2 from Arabidopsis. We showed that the cell cycle-regulated BUBR1, BUB3.1 and MAD2 proteins interacted physically with each other. Furthermore, BUBR1 and MAD2 interacted specifically at chromocenters. Following SAC activation by global defects in spindle assembly, these three interacting partners localised to unattached kinetochores. In addition, in cases of 'wait anaphase', plant SAC proteins were associated with both kinetochores and kinetochore microtubules. Unexpectedly, BUB3.1 was also found in the phragmoplast midline during the final step of cell division in plants. Conclusions/Significance: We conclude that plant BUBR1, BUB3.1 and MAD2 proteins may have the SAC protein functions conserved from yeast to humans. The association of BUB3.1 with both unattached kinetochore and phragmoplast suggests that in plant, BUB3.1 may have other roles beyond the spindle assembly checkpoint itself. Finally, this study of the SAC dynamics pinpoints uncharacterised roles of this surveillance mechanism in plant cell division