The type II bacterial CRISPR/Cas system is a novel genome-engineering technology with the ease of multiplexed gene targeting. Here, we created reporter and conditional mutant mice by coinjection of ...zygotes with Cas9 mRNA and different guide RNAs (sgRNAs) as well as DNA vectors of different sizes. Using this one-step procedure we generated mice carrying a tag or a fluorescent reporter construct in the Nanog, the Sox2, and the Oct4 gene as well as Mecp2 conditional mutant mice. In addition, using sgRNAs targeting two separate sites in the Mecp2 gene, we produced mice harboring the predicted deletions of about 700 bps. Finally, we analyzed potential off-targets of five sgRNAs in gene-modified mice and ESC lines and identified off-target mutations in only rare instances.
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•One-step generation of mice with reporters in endogenous genes•One-step generation of conditional mutant mice•Off-target analysis suggests high specificity of the CRISPR/Cas9 system
CRISPR/Cas technology is used for insertion of DNA reporter constructs into endogenous genes, derivation of conditional mutant mice and generation of deletions of defined length. Off-target mutations appear only in rare occasions.
We explore the representation of the Atlantic Meridional Overturning Circulation (AMOC) in 27 models from the CMIP6 multimodel ensemble. Comparison with RAPID and SAMBA observations suggests that the ...ensemble mean represents the AMOC strength and vertical profile reasonably well. Linear trends over the entire historical period (1850–2014) are generally neutral, but many models exhibit an AMOC peak around the 1980s. Ensemble mean AMOC decline in future (SSP) scenarios is stronger in CMIP6 than CMIP5 models. In fact, AMOC decline in CMIP6 is surprisingly insensitive to the scenario at least up to 2060. We find an emergent relationship among a majority of models between AMOC strength and 21st century AMOC decline. Constraining this relationship with RAPID observations suggests that the AMOC might decline between 6 and 8 Sv (34–45%) by 2100. A smaller group of models projects much less AMOC weakening of only up to 30%.
Plain Language Summary
The Atlantic Meridional Overturning Circulation (AMOC) is a circulation pattern in the Atlantic Ocean that is an important component of the climate system, due to its ability to redistribute and sequester heat and carbon. An accurate representation of the AMOC is a critical test for climate models and essential for building confidence in their projections. Here we investigate the AMOC in 27 climate models that contributed simulations to the Coupled Model Intercomparison Project Phase 6 (CMIP6). We find that many models reproduce the observed AMOC quite well, but there are still several models in which the AMOC is too weak or too strong. Most models suggest a slight upward trend in the AMOC from 1850 to the 1980s. Simulations representing different scenarios for future socioeconomic development suggest a stronger AMOC decline compared to previous assessments. Using direct measurements of the AMOC since 2004 and an emerging across‐model relationship between AMOC decline in the 21st century and their present‐day mean state, we find that the majority of CMIP6 models point to an end of century AMOC weakening of 34–45% of its present‐day strength. A smaller group of models projects much less weakening of only up to 30% of its present state.
Key Points
AMOC mean strength is well reproduced by the CMIP6 multimodel mean, but large model spread persists
Projected AMOC decline by the end of the 21st century shows weak dependence on the SSP scenarios
An emergent constraint between AMOC strength and projected decline suggests possible AMOC decline between 34% and 45% by 2100
Conflicting perspectives exist regarding the application of the technology acceptance model (TAM) and the theory of planned behavior (TPB) to the study of technology acceptance behavior. The present ...study addressed the controversy by evaluating and comparing the predictive power of the two theories in a specific context, which was to measure students' intentions to use a wiki for group work and their behaviors in doing so. A total of 174 students from a university in Hong Kong participated in the study. Three hypothesized models were examined using factor-based partial least squares structural equation modeling (PLS-SEM), which can account for measurement errors and is thus more robust than regression-based PLS-SEM. The results likely rebut the view that the TPB is inferior to the TAM. Moreover, this research highlighted the importance of social influences on collaborative e-learning.
Chimeric antigen receptor （CAR） T cell therapy is a promising approach to treat cancer, such as B-cell ma- lignancy 1. However, the current standard treatment requires autologous adoptive cell ...transfer, which is ex- pensive and time-consuming. For newborn and elder patients, it is often difficult to obtain enough T ceils with good quality to generate patient-specific CAR-T cells. To make CAR-T therapy more accessible, it is highly desirable to develop an allogeneic adoptive transfer strategy, in which universal CAR-T cells derived from T cells from healthy donors can be applied to treat multiple patients.
Since its commercial introduction three-quarters of a century ago, fluid catalytic cracking has been one of the most important conversion processes in the petroleum industry. In this process, porous ...composites composed of zeolite and clay crack the heavy fractions in crude oil into transportation fuel and petrochemical feedstocks. Yet, over time the catalytic activity of these composite particles decreases. Here, we report on ptychographic tomography, diffraction, and fluorescence tomography, as well as electron microscopy measurements, which elucidate the structural changes that lead to catalyst deactivation. In combination, these measurements reveal zeolite amorphization and distinct structural changes on the particle exterior as the driving forces behind catalyst deactivation. Amorphization of zeolites, in particular, close to the particle exterior, results in a reduction of catalytic capacity. A concretion of the outermost particle layer into a dense amorphous silica-alumina shell further reduces the mass transport to the active sites within the composite.Catalyst deactivation in fluid catalytic cracking processes is unavoidably associated with structural changes. Here, the authors visualize the deactivation of zeolite catalysts by ptychography and other imaging techniques, showing pronounced amorphization of the outer layer of the catalyst particles.
The directed activation of carbon-hydrogen bonds (C-H) is important in the development of synthetically useful reactions, owing to the proximity-induced reactivity and selectivity that is enabled by ...coordinating functional groups. Palladium-catalysed non-directed C-H activation could potentially enable further useful reactions, because it can reach more distant sites and be applied to substrates that do not contain appropriate directing groups; however, its development has faced substantial challenges associated with the lack of sufficiently active palladium catalysts. Currently used palladium catalysts are reactive only with electron-rich arenes, unless an excess of arene is used, which limits synthetic applications. Here we report a 2-pyridone ligand that binds to palladium and accelerates non-directed C-H functionalization with arene as the limiting reagent. This protocol is compatible with a broad range of aromatic substrates and we demonstrate direct functionalization of advanced synthetic intermediates, drug molecules and natural products that cannot be used in excessive quantities. We also developed C-H olefination and carboxylation protocols, demonstrating the applicability of our methodology to other transformations. The site selectivity in these transformations is governed by a combination of steric and electronic effects, with the pyridone ligand enhancing the influence of sterics on the selectivity, thus providing complementary selectivity to directed C-H functionalization.
We report here a catalytic method for the modular ring expansion of cyclic aliphatic alcohols. In this work, proton-coupled electron transfer activation of an allylic alcohol substrate affords an ...alkoxy radical intermediate that undergoes subsequent C–C bond cleavage to furnish an enone and a tethered alkyl radical. Recombination of this alkyl radical with the revealed olefin acceptor in turn produces a ring-expanded ketone product. The regioselectivity of this C–C bond-forming event can be reliably controlled via substituents on the olefin substrate, providing a means to convert a simple N-membered ring substrate to either n+1 or n+2 ring adducts in a selective fashion.
The ten-eleven translocation (Tet) family of methylcytosine dioxygenases catalyze oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) and promote DNA demethylation. Despite the ...abundance of 5hmC and Tet proteins in the brain, little is known about the functions of the neuronal Tet enzymes. Here, we analyzed Tet1 knockout mice (Tet1KO) and found downregulation of multiple neuronal activity-regulated genes, including Npas4, c-Fos, and Arc. Furthermore, Tet1KO animals exhibited abnormal hippocampal long-term depression and impaired memory extinction. Analysis of the key regulatory gene, Npas4, indicated that its promoter region, containing multiple CpG dinucleotides, is hypermethylated in both naive Tet1KO mice and after extinction training. Such hypermethylation may account for the diminished expression of Npas4 itself and its downstream targets, impairing transcriptional programs underlying cognitive processes. In summary, we show that neuronal Tet1 regulates normal DNA methylation levels, expression of activity-regulated genes, synaptic plasticity, and memory extinction.
•Tet1KO mice display memory extinction impairment•Tet1KO mice exhibit abnormally enhanced hippocampal LTD•Npas4, c-Fos, Arc are downregulated in Tet1KO cortex and hippocampus•Npas4 promoter is hypermethylated in Tet1KO cortex and hippocampus
Rudenko et al. report that Tet1 loss impairs hippocampal LTD and memory extinction. They find downregulation of activity-regulated genes in Tet1KO mice and hypermethylation of Npas4 promoter in naive Tet1KO mice and after memory extinction. Thus, Tet1 regulates activity-regulated genes and memory extinction.
During cellular reprogramming, only a small fraction of cells become induced pluripotent stem cells (iPSCs). Previous analyses of gene expression during reprogramming were based on populations of ...cells, impeding single-cell level identification of reprogramming events. We utilized two gene expression technologies to profile 48 genes in single cells at various stages during the reprogramming process. Analysis of early stages revealed considerable variation in gene expression between cells in contrast to late stages. Expression of Esrrb, Utf1, Lin28, and Dppa2 is a better predictor for cells to progress into iPSCs than expression of the previously suggested reprogramming markers Fbxo15, Fgf4, and Oct4. Stochastic gene expression early in reprogramming is followed by a late hierarchical phase with Sox2 being the upstream factor in a gene expression hierarchy. Finally, downstream factors derived from the late phase, which do not include Oct4, Sox2, Klf4, c-Myc, and Nanog, can activate the pluripotency circuitry.
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► Single cells exhibit high variation in gene expression early in reprogramming ► Esrrb, Utf1, Lin28, and Dppa2 mark cells with high probability to become iPSCs ► Activation of the Sox2 locus initiates a late hierarchical phase of gene expression ► Generation of iPSCs without Oct4, Sox2, Klf4, c-Myc, and Nanog
Jaenisch and colleagues use single-cell analyses to circumvent the challenges associated with studying the minority of cells in a population that reprogram to pluripotency, and they uncover mechanistic insights to the reprogramming process.
Abstract
The impact of nonequilibrium environment effects on the accuracy of quantum parameter estimation is investigated, and it is found that these effects can significantly affect estimation ...accuracy. Using an individual estimation strategy reveals that the nonequilibrium effects consistently enhance accuracy, regardless of the coupling strength between the probe and its environment. In contrast, weak memory effects undermine estimation accuracy. When employing a multi‐parameter simultaneous estimation strategy, it is observed that the nonequilibrium effects consistently improve the advantages of simultaneous estimation, as analyzed by the ratio of total variances between the two estimation scenarios. However, the memory effects on these advantages depend on the coupling strength between the qubit and the environment. These findings suggest that appropriate parameters of a nonequilibrium environment can increase the quantum Fisher information (QFI), thereby enhancing the accuracy of quantum parameter estimation. These significant results are essential for improving parameter estimation accuracy in quantum systems interacting with nonequilibrium environments.
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