Binding and distribution regularity of water molecules in high-moisture textured Antarctic krill (Euphausia superba) meat.

High-moisture extrusion technique with the advantage of high efficiency and low energy consumption is a promising strategy for processing Antarctic krill meat. Consequently, this study aimed to prepare high-moisture textured Antarctic krill meat (HMTAKM) with a rich fiber structure at different water contents (53 %, 57 %, and 61 %) and to reveal the binding and distribution regularity of water molecules, which is closely related to the fiber structure of HMTAKM and has been less studied. The hydrogen-bond network results indicated the presence of at least two or more types of water molecules with different hydrogen bonds. Increasing the water content of HMTAKM promoted the formation of hydrogen bonds between the water molecules and protein molecules, leading to the transition of the ß-sheet to the α-helix. These findings offer a novel viable processing technique for Antarctic krill and a new understanding of the fiber formation of high-moisture textured proteins.

Evaluation of the clinical application value of artificial intelligence in diagnosing head and neck aneurysms.

OBJECTIVE: To evaluate the performance of a semi-automated artificial intelligence (AI) software program (CerebralDoc® system) in aneurysm detection and morphological measurement. METHODS: In this study, 354 cases of computed tomographic angiography (CTA) were retrospectively collected in our hospital. Among them, 280 cases were diagnosed with aneurysms by either digital subtraction angiography (DSA) and CTA (DSA group, n = 102), or CTA-only (non-DSA group, n = 178). The presence or absence of aneurysms, as well as their location and related morphological features determined by AI were evaluated using DSA and radiologist findings. Besides, post-processing image quality from AI and radiologists were also rated and compared. RESULTS: In the DSA group, AI achieved a sensitivity of 88.24% and an accuracy of 81.97%, whereas radiologists achieved a sensitivity of 95.10% and an accuracy of 84.43%, using DSA results as the gold standard. The AI in the non-DSA group achieved 81.46% sensitivity and 76.29% accuracy, as per the radiologists' findings. The comparison of position consistency results showed better performance under loose criteria than strict criteria. In terms of morphological characteristics, both the DSA and the non-DSA groups agreed well with the diagnostic results for neck width and maximum diameter, demonstrating excellent ICC reliability exceeding 0.80. The AI-generated images exhibited superior quality compared to the standard software for post-processing, while also demonstrating a significantly reduced processing time. CONCLUSIONS: The AI-based aneurysm detection rate demonstrates a commendable performance, while the extracted morphological parameters exhibit a remarkable consistency with those assessed by radiologists, thereby showcasing significant potential for clinical application.

Quantifying the historical and future heat-related mortality above the heat alert thresholds of the inaugural Chinese national heat-health action plan.

BACKGROUND: China published its inaugural national heat-health action plan (HHAP) in 2023, but the mortality burden associated with temperatures exceeding the heat alert thresholds specified by this HHAP (maximum temperatures >35, 37, or 40 °C) remains unknown. We aimed to estimate the historical and future mortality burden associated with temperatures above the heat alert thresholds of the Chinese national HHAP. METHODS: We conducted time-series analyses to estimate the mortality burden associated with temperatures exceeding the three heat alert thresholds from 2016 to 2019 in Jiangsu Province (including 13 cities, population ∼80.7 million), China. A quasi-Poisson regression in conjunction with a distributed lag non-linear model was used to estimate the dose-response association between maximum temperature and mortality risk from 2016 to 2019, adjusting for potential covariates. We then projected the future mortality burden associated with temperatures exceeding these thresholds under three distinct levels of greenhouse gas (GHG) emission scenarios via scenario shared socioeconomic pathways [SSP] 1-2.6 (low), SSP2-4.5 (intermediate), and SSP5-8.5 (high), respectively, by assuming that there will be no adaptation to heat. Climate scenarios derived from the General Circulation Model (GCM) under the Coupled Model Intercomparison Project Phase 6 (CMIP6) were used. RESULTS: From 2016 to 2019, temperatures above 35 °C were associated with 0.51% of mortality, including 0.40% associated with 35 °C-37 °C and 0.11% associated with >37 °C. Heat-related mortality risk was most prominent in those who were single/divorced/widowed and had <10 years of education. Under SSP2-4.5, compared with the 2020s, the excess mortality associated with >37 °C would increase by 1.4 times in the 2050s and 1.7 times in the 2090s. Under SSP5-8.5, the annual number of days with maximum temperature >37 °C would approximately double every 20 years (67 days annually in the 2090s). Consequently, compared with the 2020s, the excess mortality associated with >37 °C would increase by 2.8 times in the 2050s and 18.4 times in the 2090s. CONCLUSION: Significant mortality risk is associated with temperatures above the lowest heat alert threshold of the Chinese national HHAP (35 °C). If the high GHG emission scenario occurred, the annual number of days and excess mortality associated with maximum temperatures >37 °C would largely increase in the coming decades.

Assessment of LINC-PINT genetic polymorphisms and esophageal squamous cell carcinoma risk in the Hainan Han population.

OBJECTIVES: Esophageal squamous cell carcinoma (ESCC) is a malignant tumor with high incidence and mortality rates worldwide. This study aimed to investigate the correlation between LINC-PINT polymorphisms and ESCC risk in the Hainan Han population. METHODS: A total of 391 patients with ESCC and 452 healthy controls were enrolled to evaluate the effect of LINC-PINT SNPs (single nucleotide polymorphisms) on ESCC susceptibility. Associations were evaluated by calculating odds ratios (OR) and 95% confidence intervals (CIs). Multifactor dimensionality reduction analysis was performed to explore the association between SNP-SNP interactions and ESCC susceptibility. We further determined the correlation between clinical indicators and SNP in patients with ESCC. RESULTS: Our study showed that rs157916 (OR 0.63, p = 0.011) and rs157928 (OR 0.80, p = 0.021) were associated with a decreased risk of ESCC. Stratified analysis indicated that rs157916 could decrease the risk of ESCC in people aged >64 years, in males, and non-drinkers (OR 0.58, p = 0.042; OR 0.58, p = 0.010; OR 0.62, p = 0.025, respectively). Rs16873842 was related to a decreased risk of ESCC in males (OR 0.70, p = 0.015). Rs7801029 was associated with ESCC risk in females (OR 0.39, p = 0.033) and non-drinkers (OR 0.68, p = 0.040). Rs7781295 decreased the ESCC risk in smokers (OR 0.58, p = 0.046) and drinkers (OR 0.58, p = 0.046). In addition, rs157928 played a protective role in ESCC risk in females (OR 0.39, p = 0.033) and non-smokers (OR 0.32, p = 0.006). Additionally, the best predictive model for ESCC was a combination of rs157916, rs16873842, rs7801029, rs7781295, rs28662387, and rs157928. CONCLUSION: Our study revealed that LINC-PINT polymorphisms were associated with ESCC risk.

Targeted degradation of LRG1 to attenuate renal fibrosis.

Leucine-rich α-2 glycoprotein 1 (LRG1), a secreted glycoprotein, has been identified as significantly upregulated in renal fibrosis, potentially exacerbating the condition by enhancing TGF-ß-Smad3-dependent signaling pathways. Herein, utilizing our developed LRG1-targeting peptide for LRG1 recruitment and lenalidomide for E3 ubiquitin ligase engagement, we developed an advanced proteolysis targeting chimera, ETTAC-2, specifically designed for LRG1 degradation. Our cellular degradation assays validated that ETTAC-2 effectively degraded LRG1 through a proteasome-dependent mechanism, achieving half-maximal degradation at a concentration of 8.38 µM. Furthermore, anti-fibrotic experiments conducted both in vitro and in vivo revealed that ETTAC-2 efficiently induced LRG1 degradation in fibrotic kidneys. This action effectively inhibited the TGF-ß-Smad3 signaling pathway and diminished the secretion of fibrosis-associated proteins, consequently attenuating the progression of renal fibrosis. Our study highlights the pivotal role of LRG1 in renal fibrosis and positions ETTAC-2 as a promising therapeutic candidate for targeted LRG1 intervention.

A new fusion strategy for rapid strain differentiation based on MALDI-TOF MS and Raman spectra.

Typing of bacterial subspecies is urgently needed for the diagnosis and efficient treatment during disease outbreaks. Physicochemical spectroscopy can provide a rapid analysis but its identification accuracy is still far from satisfactory. Herein, a novel feature-extractor-based fusion-assisted machine learning strategy has been developed for high accuracy and rapid strain differentiation using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and Raman spectroscopy. Based on this fusion approach, rapid and reliable identification and analysis can be performed within 24 hours. Validation on a panel of important pathogens comprising Staphylococcus aureus, Klebsiella pneumoniae, Escherichia coli, and Acinetobacter baumannii showed that the identification accuracies of k-nearest neighbors (KNNs), support vector machines (SVMs) and artificial neural networks (ANNs) were 100%. In particular, when benchmarked against a MALDI-TOF MS spectral dataset, the new approach improved the identification accuracy of Acinetobacter baumannii from 87.67% to 100%. This work demonstrates the effectiveness of combining MALDI-TOF MS and Raman spectroscopy fusion data in pathogenic bacterial subtyping.

Ferroptosis contributes to lead-induced cochlear spiral ganglion neurons injury.

The underlying mechanisms of lead exposure-induced cochlear spiral ganglion neurons (SGNs) injury are not yet clear. This study explored whether ferroptosis is involved in lead-induced SGNs injury and investigated the mechanism of lead-induced iron overload in SGNs. A primary culture cell model of lead acetate-induced SGNs damage was established. The changes in levels of iron ions, reactive oxygen species, lipid peroxides, and glutathione in SGNs were measured after lead acetate intervention and ferroptosis inhibitors pre-treatment. The morphology of mitochondria was also observed, and the expression of ferroptosis marker genes glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11), as well as iron metabolism-related proteins transferrin receptor protein 1 (TFR1), nuclear receptor coactivator 4 (NCOA4), heme oxygenase-1 (HO-1), and ferroportin (FPN) were detected. Results showed that lead acetate exposure induced SGNs injury in a time- and dose-dependent manner. Intracellular iron accumulation, increased levels of reactive oxygen species and lipid peroxide with decreased level of antioxidant capacity were occurred in SGNs after lead exposure. Meanwhile, decreased expressions of GPX4 and SLC7A11 and increased expressions of iron metabolism-related proteins (TFR1, NCOA4, and HO-1) were also found. Lead acetate intervention also caused mitochondrial shrinkage with blurred and fragmented morphology. Pre-treatment with ferroptosis inhibitors (Fer-1 and DFOM) significantly ameliorated lead-induced SGNs injury. In summary, lead exposure can induce ferroptosis in SGNs, the antioxidant defense system and iron metabolism disorder are involved in lead-induced SGNs ferroptosis. Thus, inhibiting ferroptosis may be a new strategy for preventing and treating lead exposure-related hearing loss.

Genome-wide analysis of the AP2/ERF gene family in Pennisetum glaucum and the negative role of PgRAV_01 in drought tolerance.

APETALA2/ethylene-responsive (AP2/ERF) plays crucial roles in resisting diverse stresses and in regulating plant growth and development. However, little is known regarding the structure and function of the AP2/ERF genes in pearl millet (Pennisetum glaucum). The AP2/ERF gene family may be involved in the development and maintenance of P. glaucum resilience to abiotic stresses, central to its role as a vital forage and cereal crop. In this study, PgAP2/ERF family members were identified and comprehensive bioinformatics analyses were performed, including determination of phylogenetic relationships, gene structures, conserved motifs, chromosomal localization, gene duplication, expression pattern, protein interaction network, and functional characterization of PgRAV_01 (Related to ABI3/VP1). In total, 78 PgAP2/ERF members were identified in the P. glaucum genome and classified into five subfamilies: AP2, ERF, DREB, RAV, and soloist. Members within the same clade of the PgAP2/ERF family showed similar gene structures and motif compositions. Six duplication events were identified in the PgAP2/ERF family; calculation of Ka/Ks values showed that purification selection dominated the evolution of PgAP2/ERFs. Subsequently, a potential interaction network of PgAP2/ERFs was generated to predict the interaction relationships. Additionally, abiotic stress expression analysis showed that most PgAP2/ERFs were induced in response to drought and heat stresses. Furthermore, overexpression of PgRAV_01 negatively regulated drought tolerance in Nicotiana benthamiana by reducing its antioxidant capacity and osmotic adjustment. Taken together, these results provide valuable insights into the characteristics and functions of PgAP2/ERF genes, with implications for abiotic stress tolerance, and will ultimately contribute to the genetic improvement of cereal crop breeding.

Advanced precipitation peak offsets middle growing-season drought in impacting grassland C sink.

Redistribution of precipitation across seasons is a widespread phenomenon affecting dryland ecosystems globally. However, the impacts of shifting seasonal precipitation patterns on carbon (C) cycling and sequestration in dryland ecosystems remain poorly understood. In this study, we conducted a 10-yr (2013-2022) field manipulative experiment that altered the timing of growing-season precipitation peaks in a semi-arid grassland. We found that the delayed precipitation peak suppressed plant growth and thus reduced gross ecosystem productivity, ecosystem respiration, and net ecosystem productivity due to middle growing-season water stress. Surprisingly, shifting more precipitation to the early growing season can advance plant development, increase the dominance of drought-tolerant forbs, and thus compensate for the negative impacts of middle growing-season water stress on ecosystem C cycling, leading to a neutral change in grassland C sink. Our findings indicate that greater precipitation and plant development in spring could act as a crucial mechanism, maintaining plant growth and stabilizing ecosystem C sink. This underscores the urgent need to incorporate precipitation seasonality into Earth system models, which is crucial for improving projections of terrestrial C cycling and sequestration under future climate change scenarios.

CXCR4 regulates macrophage M1 polarization by altering glycolysis to promote prostate fibrosis.

BACKGROUND: C-X-C receptor 4(CXCR4) is widely considered to be a highly conserved G protein-coupled receptor, widely involved in the pathophysiological processes in the human body, including fibrosis. However, its role in regulating macrophage-related inflammation in the fibrotic process of prostatitis has not been confirmed. Here, we aim to describe the role of CXCR4 in modulating macrophage M1 polarization through glycolysis in the development of prostatitis fibrosis. METHODS: Use inducible experimental chronic prostatitis as a model of prostatic fibrosis. Reduce CXCR4 expression in immortalized bone marrow-derived macrophages using lentivirus. In the fibrotic mouse model, use adenovirus carrying CXCR4 agonists to detect the silencing of CXCR4 and assess the in vivo effects. RESULTS: In this study, we demonstrated that reducing CXCR4 expression during LPS treatment of macrophages can alleviate M1 polarization. Silencing CXCR4 can inhibit glycolytic metabolism, enhance mitochondrial function, and promote macrophage transition from M1 to M2. Additionally, in vivo functional experiments using AAV carrying CXCR4 showed that blocking CXCR4 in experimental autoimmune prostatitis (EAP) can alleviate inflammation and experimental prostate fibrosis development. Mechanistically, CXCR4, a chemokine receptor, when silenced, weakens the PI3K/AKT/mTOR pathway as its downstream signal, reducing c-MYC expression. PFKFB3, a key enzyme involved in glucose metabolism, is a target gene of c-MYC, thus impacting macrophage polarization and glycolytic metabolism processes.

Construction and validation of a nomogram model for predicting different sites of ankle pain in runners with chronic ankle instability.

This study aimed to establish a risk prediction nomogram model for anterolateral, mediolateral, and posterolateral ankle pain in runners with chronic ankle instability (CAI) and analyse the potential risk factors for pain at different ankle sites. Thirty recreational runners with CAI who reported ankle pain in the anterolateral, mediolateral, or posterolateral regions were recruited for this study. Kinematic, kinetic, and electromyographic data during running were collected using motion capture system, 3-D force platform, and surface electromyography system. These data were used to generate a dynamic nomogram. The results showed that anterolateral ankle pain in runners with CAI may be caused by insufficient gastrocnemius muscle strength (OR 0.85, 95% CI 0.73-0.97), excessive ground reaction force (GRF, OR 2.64, 95% CI 1.25-6.22), and an increased percentage of ankle energy absorption (OR 9.11, 95% CI 1.50-77.79). Mediolateral ankle pain might be contributed by greater ankle inversion angle (OR 1.08, 95% CI 1.01-1.00) and GRF (OR 2.13, 95% CI 1.17-4.31). Moreover, posterolateral ankle pain was predicted by increased ankle adduction angle (OR 1.06, 95% CI 1.00-1.12), increased GRF (OR 2.16, 95% CI 1.07-4.80), and decreased dynamic stability (OR 0.20, 95% CI 0.05-0.68). To prevent ankle pain, runners with CAI should be encouraged to focus on improving the neuroreceptor sensitivity of the gastrocnemius muscles, and retraining their energy absorption patterns.

Machine Learning-Driven discovery of immunogenic cell Death-Related biomarkers and molecular classification for diabetic ulcers.

In this study, we redefine the diagnostic landscape of diabetic ulcers (DUs), a major diabetes complication. Our research uncovers new biomarkers linked to immunogenic cell death (ICD) in DUs by utilizing RNA-sequencing data of Gene Expression Omnibus (GEO) analysis combined with a comprehensive database interrogation. Employing a random forest algorithm, we have developed a diagnostic model that demonstrates improved accuracy in distinguishing DUs from normal tissue, with satisfactory results from ROC analysis. Beyond mere diagnosis, our model categorizes DUs into novel molecular classifications, which may enhance our comprehension of their underlying pathophysiology. This study bridges the gap between molecular insights and clinical practice. It sets the stage for transformative strategies in DUs management, marking a significant step forward in personalized medicine for diabetic patients.

Patterns of nitrate load variability under surface water-groundwater interactions in agriculturally intensive valley watersheds.

Nitrate pollution is a significant environmental issue closely related to human activities, complicated hydrological interactions and nitrate fate in the valley watershed strongly affects nitrate load in hydrological systems. In this study, a nitrate reactive transport model by coupling SWAT-MODFLOW-RT3D between surface water and groundwater interactions at the watershed scale was developed, which was used to reproduce the interaction between surface water and groundwater in the basin from 2016 to 2019 and to reveal the nitrogen transformation process and the evolving trend of nitrate load within the hydrological system of the valley watershed. The results showed that the basin exhibited groundwater recharge to surface water in 2016-2019, particularly in the northwestern and northeastern mountainous regions of the valley watershed and the southern Beishan Reservoir vicinity. Groundwater recharge to surface water declined by 20.17 % from 2016 to 2019 due to precipitation. Nitrate loads in the hydrologic system of the watershed are primarily derived from human activities (including fertilizer application from agricultural activities and residential wastewater discharges) and the nitrogen cycle. Nitrate loads in surface water declined 16.05 % from 2016 to 2019. Nitrate levels are higher in agricultural farming and residential areas on the eastern and northern sides of the watershed. Additionally, hydrological interactions are usually accompanied by material accumulation and environmental changes. Nitrate levels tend to rise with converging water flows, a process that becomes more pronounced during precipitation events and cropping seasons in agriculturally intensive valley watersheds. However, environmental changes alter nitrogen transformation processes. Nitrogen fixation, nitrification, and ammonification intensify nitrogen inputs during river pooling, enhancing nitrogen cycling fluxes and elevating nitrate loads. These processes are further enhanced during groundwater recharge to surface water, leading to evaluated nitrate load. Enhanced denitrification, dissimilatory nitrate reduction to ammonium (DNRA), anaerobic ammonia oxidation, and assimilation promote the nitrogen export from the system and reduce the nitrate load during surface water recharge to groundwater.

Melting and Rapid Solidification of Lunar Regolith Particles Returned by Chang'E-5 Mission.

Melting and solidification of lunar regolith are pivotal for comprehending the evolutionary dynamics of lunar volcanism, geology, and impact history. Additionally, insights gained from these processes can contribute to the advancement of in situ resource utilization technologies, for instance additive manufacturing and resource extraction systems. Herein, we conduct the direct observation of the melting and rapid solidification of lunar particles returned by the Chang'E 5 mission. The melting temperature and melting sequence were obtained. Bubble generation, growth, and release were clearly observed, with a maximum bubble diameter of 5 µm, which is supposed to be according to the release of volatiles that embedded in the particles. During the solidification process, evident crystallization occurred with incremental crystal growth rate approximately of 27 nm/s. Scanning electron microscopy and energy-dispersive x-ray spectroscopy results verified that the Fe-rich mineral crystalizes first. These results would improve the understanding of the evolution of lunar volcanism, geology, and impact history.

Applications of Digital Enzyme-Linked Immunosorbent Assays in Ophthalmology.

Digital enzyme-linked immunosorbent assays (dELISAs) very sensitively detect biomarkers that cannot be measured using traditional methods. The molecules are confined within a small volume, their counts accurately computed, and the results rapidly delivered. Digital ELISAs find many applications. In recent years, such ELISAs have become increasingly used to aid ophthalmological diagnoses and treatments, and have revolutionized the field. This article reviews the applications of dELISAs in clinical practice, especially in the sphere of ophthalmology.

Transient Phase-Mediated Li+ Transportation in the Lithium Lanthanum Titanate Solid-State Electrolyte.

The lithium lanthanum titanium oxide (LLTO) perovskite is one type of superior lithium (Li)-ion conductor that is of great interest as a solid-state electrolyte for all-solid-state lithium batteries. Structural defects and impurity phases formed during the synthesis of LLTO largely affect its Li-ion conductivity, yet the underlying Li+ diffusion mechanism at the atomic scale is still under scrutiny. Herein, we use aberration-corrected transmission electron microscopy to perform a thorough structural characterization of the LLTO ceramic pellet. We reveal a prevalent transient phase transition of (La, Ti)2O3 existing at the antiphase boundaries between single-crystalline LLTO domains. This transient phase exhibits a specific crystal orientation with the LLTO phase and shows a gradual structural transition to a tetragonal LLTO structure, which enables detailed crystallographic analysis to correlate their formation to the sintering process of LLTO powders into ceramic pellets. We also find that Li diffusion is retarded by this phase and correlated with the excess amount of La, which is corroborated by the theoretical evaluation of the atomistic mechanisms of Li diffusion across this phase.

Research Progress on Photoperiod Gene Regulation of Heading Date in Rice.

Heading date is a critical physiological process in rice that is influenced by both genetic and environmental factors. The photoperiodic pathway is a primary regulatory mechanism for rice heading, with key florigen genes Hd3a (Heading date 3a) and RFT1 (RICE FLOWERING LOCUS T1) playing central roles. Upstream regulatory pathways, including Hd1 and Ehd1, also significantly impact this process. This review aims to provide a comprehensive examination of the localization, cloning, and functional roles of photoperiodic pathway-related genes in rice, and to explore the interactions among these genes as well as their pleiotropic effects on heading date. We systematically review recent advancements in the identification and functional analysis of genes involved in the photoperiodic pathway. We also discuss the molecular mechanisms underlying rice heading date variation and highlight the intricate interactions between key regulatory genes. Significant progress has been made in understanding the molecular mechanisms of heading date regulation through the cloning and functional analysis of photoperiod-regulating genes. However, the regulation of heading date remains complex, and many underlying mechanisms are not yet fully elucidated. This review consolidates current knowledge on the photoperiodic regulation of heading date in rice, emphasizing novel findings and gaps in the research. It highlights the need for further exploration of the interactions among flowering-related genes and their response to environmental signals. Despite advances, the full regulatory network of heading date remains unclear. Further research is needed to elucidate the intricate gene interactions, transcriptional and post-transcriptional regulatory mechanisms, and the role of epigenetic factors such as histone methylation in flowering time regulation. This review provides a detailed overview of the current understanding of photoperiodic pathway genes in rice, setting the stage for future research to address existing gaps and improve our knowledge of rice flowering regulation.

Exclusive generation of a superoxide radical by a porous aromatic framework for fast photocatalytic decontamination of mustard gas simulant in room air.

Mustard gas and other chemical warfare agents (CWAs) are a global threat to public security, arising from unpredictable emergencies and chemical spill accidents. So far, photocatalysts such as metal clusters, polyoxometalates and porous solids have been exploited for oxidative degradation of mustard gas, commonly with 1O2 as reactive species. However, the production of 1O2 is oxygen-dependent and requires a high oxygen concentration to sustain the detoxication process. For safety and operation process considerations, it is always preferable to rapidly detoxify dangerous chemicals in the atmosphere of room air. In this work, a porous aromatic framework, PAF-68, was synthesized as a metal-free photocatalyst. In the presence of PAF-68, fast detoxication occurred in typical room air atmosphere. The half-life (t 1/2) for the complete conversion of mustard gas simulant to nontoxic product in room air was only 1.7 min, which is comparable to the performance in pure oxygen, surpassing that of any other porous photocatalysts. It was found that ˙O2 - rather than 1O2 is the predominant reactive species initiated by PAF-68 for mustard gas detoxication. Unlike the formation of 1O2 which prefers the environment of pure oxygen, generation of the ˙O2 - is an oxygen-independent process. It is suggested that amorphous PAFs possess low exciton binding energy and long decay lifetime, which facilitate the generation of ˙O2 -, and this offers a general design strategy to detoxifying chemical warfare agents under real-world conditions.

Construction and validation of co-expression vector for rice alpha tubulin and microtubule associated protein respectively fused with fluorescent proteins.

Microtubule (MT) consists of α-tubulin and ß-tubulin. The dynamic instability regulated by various microtubule associated proteins (MAPs) is essential for MT functions. To analyze the interaction between tubulin/MT and MAP in vivo, we usually need tubulin and MAP co-expressed. Here, we constructed a dual-transgene vector expressing rice (Oryza sativa) α-tubulin and MAP simultaneously. To construct this vector, plant expression vector pCambia1301 was used as the plasmid backbone and Gibson assembly cloning technology was used. We first fused and cloned the GFP fragment, α-tubulin open reading frame (ORF), and NOS terminator into the vector pCambia1301 to construct the p35S::GFP-α-tubulin vector that expressed GFP-α-tubulin fusion protein. Subsequently, we fused and cloned the CaMV 35S promoter, mCherry fragment, and NOS terminator into the p35S::GFP-α-tubulin vector to generate the universal dual-transgene expression vector (p35S::GFP-α-tubulin-p35S::mCherry vector). With the p35S::GFP-α-tubulin-p35S::mCherry vector, MAP ORF can be cloned into the site of 5' or 3' terminus of mCherry to co-express GFP-α-tubulin and MAP-mCherry/mCherry-MAP. To validate the availability and universality of the dual-transgene expression vector, a series of putative rice MAP genes including GL7, OsKCBP, OsCLASP, and OsMOR1 were cloned into the vector respectively, transformed into Agrobacterium tumefaciens strain, and expressed in Nicotiana benthamiana leaves. The results indicated that all of the MAPs were co-expressed with α-tubulin and localized to MTs, validating the availability and universality of the vector and that GL7, OsKCBP, OsCLASP, and OsMOR1 might be MAPs. The application of the co-expression vector constructed by us would facilitate studies on the interaction between tubulin/MT and MAP in tobacco transient expression systems or transgenic rice.