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"Wang, Xue-Chen"
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Functional pavements : proceedings of the 6th Chinese-European Workshop on Functional Pavement Design (CEW 2020), Nanjing, China, 18-21 October 2020
\"Functional Pavements is a collection of papers presented at the 6th Chinese-European Workshop (CEW) on Functional Pavement Design (Nanjing, China, October 18-21, 2020). The focus of the CEW series is on field tests, laboratory test methods and advanced analysis techniques, and cover analysis, material development and production, experimental characterization, design and construction of pavements. The main areas covered by the book include: Asphalt binders for flexible pavements Asphalt mixture evaluation and performance Pavement construction and maintenance Pavement Surface Properties and Vehicle Interaction Cementitious materials for rigid pavements Pavement geotechnics and environment Functional Pavements aims at contributing to the establishment of a new generation of pavement design methodologies in which rational mechanics principles, advanced constitutive models and advanced material characterization techniques shall constitute the backbone of the design process. The book will be much of interest to professionals, academics and practitioners in pavement engineering and related disciplines as it should assist them in providing improved road pavement infrastructure to their stakeholders.\"-- Provided by publisher.
Book
Egg cell-specific promoter-controlled CRISPR/Cas9 efficiently generates homozygous mutants for multiple target genes in Arabidopsis in a single generation
Arabidopsis mutants produced by constitutive overexpression of the CRISPR/Cas9 genome editing system are usually mosaics in the T1 generation. In this study, we used egg cell-specific promoters to drive the expression of Cas9 and obtained non-mosaic T1 mutants for multiple target genes with high efficiency. Comparisons of 12 combinations of eight promoters and two terminators found that the efficiency of the egg cell-specific promoter-controlled CRISPR/Cas9 system depended on the presence of a suitable terminator, and the composite promoter generated by fusing two egg cell-specific promoters resulted in much higher efficiency of mutation in the T1 generation compared with the single promoters.
Journal Article
Prime editing efficiently generates W542L and S621I double mutations in two ALS genes in maize
Prime editing is a novel and universal CRISPR/Cas-derived precision genome-editing technology that has been recently developed. However, low efficiency of prime editing has been shown in transgenic rice lines. We hypothesize that enhancing pegRNA expression could improve prime-editing efficiency. In this report, we describe two strategies for enhancing pegRNA expression. We construct a prime editing vector harboring two pegRNA variants for W542L and S621I double mutations in
ZmALS1
and
ZmALS2
. Compared with previous reports in rice, we achieve much higher prime-editing efficiency in maize. Our results are inspiring and provide a direction for the optimization of plant prime editors.
Journal Article
A CRISPR/Cas9 toolkit for multiplex genome editing in plants
Background
To accelerate the application of the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/ CRISPR-associated protein 9) system to a variety of plant species, a toolkit with additional plant selectable markers, more gRNA modules, and easier methods for the assembly of one or more gRNA expression cassettes is required.
Results
We developed a CRISPR/Cas9 binary vector set based on the pGreen or pCAMBIA backbone, as well as a gRNA (guide RNA) module vector set, as a toolkit for multiplex genome editing in plants. This toolkit requires no restriction enzymes besides
Bsa
I to generate final constructs harboring maize-codon optimized
Cas9
and one or more gRNAs with high efficiency in as little as one cloning step. The toolkit was validated using maize protoplasts, transgenic maize lines, and transgenic
Arabidopsis
lines and was shown to exhibit high efficiency and specificity. More importantly, using this toolkit, targeted mutations of three
Arabidopsis
genes were detected in transgenic seedlings of the T1 generation. Moreover, the multiple-gene mutations could be inherited by the next generation.
Conclusions
We developed a toolkit that facilitates transient or stable expression of the CRISPR/Cas9 system in a variety of plant species, which will facilitate plant research, as it enables high efficiency generation of mutants bearing multiple gene mutations.
Journal Article
Potential high-frequency off-target mutagenesis induced by CRISPR/Cas9 in Arabidopsis and its prevention
Key messageWe present novel observations of high-specificity SpCas9 variants, sgRNA expression strategies based on mutant sgRNA scaffold and tRNA processing system, and CRISPR/Cas9-mediated T-DNA integrations.Specificity of CRISPR/Cas9 tools has been a major concern along with the reports of their successful applications. We report unexpected observations of high frequency off-target mutagenesis induced by CRISPR/Cas9 in T1 Arabidopsis mutants although the sgRNA was predicted to have a high specificity score. We also present evidence that the off-target effects were further exacerbated in the T2 progeny. To prevent the off-target effects, we tested and optimized two strategies in Arabidopsis, including introduction of a mCherry cassette for a simple and reliable isolation of Cas9-free mutants and the use of highly specific mutant SpCas9 variants. Optimization of the mCherry vectors and subsequent validation found that fusion of tRNA with the mutant rather than the original sgRNA scaffold significantly improves editing efficiency. We then examined the editing efficiency of eight high-specificity SpCas9 variants in combination with the improved tRNA-sgRNA fusion strategy. Our results suggest that highly specific SpCas9 variants require a higher level of expression than their wild-type counterpart to maintain high editing efficiency. Additionally, we demonstrate that T-DNA can be inserted into the cleavage sites of CRISPR/Cas9 targets with high frequency. Altogether, our results suggest that in plants, continuous attention should be paid to off-target effects induced by CRISPR/Cas9 in current and subsequent generations, and that the tools optimized in this report will be useful in improving genome editing efficiency and specificity in plants and other organisms.
Journal Article
A Subclass of Gamma-Ray Burst Originating from Compact Binary Merger
Type I gamma-ray bursts (GRBs) are believed to originate from compact binary mergers usually with a duration of main emission less than 2 s. However, recent observations of GRB 211211A and GRB 230307A indicate that some merger-origin GRBs could last much longer. Since they show strikingly similar properties (indicating a common mechanism), which are different from the classic “long”-short burst (e.g., GRB 060614), we find they form an interesting subclass of type I GRBs, and we suggest to name them as type IL GRB. We find that the prompt emission of type IL GRB is composed of three episodes: (1) a precursor followed by a short quiescent (or weak emission) period, (2) a long-duration main emission, and (3) an extended emission. With this burst pattern, a good candidate, GRB 170228A, was found in the Fermi/Gamma-ray Burst Monitor archive data. Temporal and spectral analyses indeed show that GRB 170228A falls in the same group with GRB 211211A and GRB 230307A in many diagnostic figures. Thus, this burst pattern could be a good reference for rapidly identifying type IL GRBs and very helpful for conducting low-latency follow-up observation. We estimated the occurrence rate and discussed the physical origins and implications for the three emission episodes of type IL GRBs. Our analysis suggests the premerger precursor model, especially the magnetar super flare model, is more favored for type IL GRBs. More observations in multiwavelength and multimessenger are required to deepen our understanding of this subclass of GRB.
Journal Article
Spectral Evidence of Heavy Nuclei from the Neutron Star Crust in Magnetar Bursts
The crust of a neutron star (NS) provides a unique laboratory for studying matter under extreme density and magnetic field conditions that cannot be realized in terrestrial experiments. However, direct observational constraints on its composition have remained very limited. Magnetar bursts provide a promising means to probe the nuclear composition of the outer crust, as their energy release may be associated with stress-driven yielding of the crustal Coulomb lattice (including plastic deformation) and magnetic reconnection in the surrounding magnetosphere. We develop a general-purpose radiative transfer framework for a strongly magnetized electron–ion thermal plasma (MEITP) and apply it to the observed X-ray burst spectra. The spectral fits disfavor light-ion compositions and instead favor plasmas characterized by effective charge numbers around Z ∼ 37. These results provide spectral evidence for the participation of heavy nuclei in magnetar bursts, offer new observational constraints on the baryonic content and the location of the emitting fireballs, and further imply a crustal origin of the heavy ions.
Journal Article
Arabidopsis Glutathione Peroxidase Functions as Both a Redox Transducer and a Scavenger in Abscisic Acid and Drought Stress Responses
We isolated two T-DNA insertion mutants of Arabidopsis thaliana GLUTATHIONE PEROXIDASE3 (ATGPX3) that exhibited a higher rate of water loss under drought stress, higher sensitivity to H₂O₂ treatment during seed germination and seedling development, and enhanced production of H₂O₂ in guard cells. By contrast, lines engineered to overexpress ATGPX3 were less sensitive to drought stress than the wild type and displayed less transpirational water loss, which resulted in higher leaf surface temperature. The atgpx3 mutation also disrupted abscisic acid (ABA) activation of calcium channels and the expression of ABA- and stress-responsive genes. ATGPX3 physically interacted with the 2C-type protein phosphatase ABA INSENSITIVE2 (ABI2) and, to a lesser extent, with ABI1. In addition, the redox states of both ATGPX3 and ABI2 were found to be regulated by H₂O₂. The phosphatase activity of ABI2, measured in vitro, was reduced approximately fivefold by the addition of oxidized ATGPX3. The reduced form of ABI2 was converted to the oxidized form by the addition of oxidized ATGPX3 in vitro, which might mediate ABA and oxidative signaling. These results suggest that ATGPX3 might play dual and distinctive roles in H₂O₂ homeostasis, acting as a general scavenger and specifically relaying the H₂O₂ signal as an oxidative signal transducer in ABA and drought stress signaling.
Journal Article
Observation of spectral lines in the exceptional GRB 221009A
As the brightest gamma-ray burst ever observed, GRB 221009A provided a precious opportunity to explore spectral line features. In this article, we performed a comprehensive spectroscopy analysis of GRB 221009A jointly with GECAM-C and
Fermi
/GBM data to search for emission and absorption lines. For the first time we investigated the line feature throughout this GRB including the most bright part where many instruments suffered problems, and identified prominent emission lines in multiple time intervals. The central energy of the Gaussian emission line evolves from about 37 to 6 MeV, with a nearly constant ratio (about 10%) between the line width and central energy. Particularly, we find that both the central energy and the energy flux of the emission line evolve with time as a power law decay with power law index of −1 and −2, respectively. We suggest that the observed emission lines most likely origin from the blue-shifted electron positron pair annihilation 511 keV line. We find that a standard high latitude emission scenario cannot fully interpret the observation, thus we propose that the emission line comes from some dense clumps with electron positron pairs traveling together with the jet. In this scenario, we can use the emission line to directly, for the first time, measure the bulk Lorentz factor of the jet (Γ) and reveal its time evolution (i.e., Γ ∼
t
−1
) during the prompt emission. Interestingly, we find that the flux of the annihilation line in the co-moving frame keeps constant. These discoveries of the spectral line features shed new and important lights on the physics of GRB and relativistic jet.
Journal Article
Synchrotron Radiation Dominates the Extremely Bright GRB 221009A
The brightest gamma-ray burst, GRB 221009A, has spurred numerous theoretical investigations, with particular attention paid to the origins of ultrahigh-energy TeV photons during the prompt phase. However, analyzing the mechanism of radiation of photons in the ∼MeV range has been difficult because the high flux causes pileup and saturation effects in most GRB detectors. In this Letter, we present systematic modeling of the time-resolved spectra of the GRB using unsaturated data obtained from the Fermi Gamma-ray Burst Monitor (precursor) and SATech-01/GECAM-C (main emission and flare). Our approach incorporates the synchrotron radiation model, which assumes an expanding emission region with relativistic speed and a global magnetic field that decays with radius, and successfully fits such a model to the observational data. Our results indicate that the spectra of the burst are fully in accordance with a synchrotron origin from relativistic electrons accelerated at a large emission radius. The lack of thermal emission in the prompt emission spectra supports a Poynting flux–dominated jet composition.
Journal Article