Search Results (4,660)

Reverse transcription quantitative real-time PCR (RT-qPCR) is a highly efficient and sensitive technique for quantifying gene transcript levels. The accuracy of gene expression analysis depends critically on the selection of appropriate reference genes for normalization, which is essential to minimize technical variation arising from differences in RNA quality, reverse transcription efficiency, and sample handling. Schisandra chinensis is a medicinally important plant with a long history of use in traditional Chinese medicine and has gained increasing global recognition. In recent years, a growing number of studies have employed molecular biology approaches to investigate the molecular mechanisms underlying secondary metabolite biosynthesis in S. chinensis. However, systematically validated reference genes for RT-qPCR analysis in this species have not yet been established. In the present study, the expression stability of eleven candidate reference genes was evaluated across different tissues and under various abiotic stress conditions in S. chinensis using four statistical algorithms: geNorm, NormFinder, BestKeeper, and RefFinder. Comprehensive analysis revealed that PP2A15 and UBC2 were the optimal reference gene combination for leaves; UBC2 and UBC11 for stems; RPL6 and PP2A15 for roots; RPL21 and RPL6 for fruits; and RPL6 and UBC11 as the best-performing pair across all tissue types. Under abiotic stress conditions, UBC11 and UBC2 exhibited the highest stability in both leaves and roots under salt stress; UBC2 and GPN1 proved most stable under alkaline stress; UBC2 and RPL6 were identified as the most suitable combination under drought stress; and UBC2 and UBQ12 demonstrated consistently stable expression across all three abiotic stress treatments. The reliability of these reference gene combinations was further validated by examining the expression profiles of three target genes. Collectively, these findings establish a validated reference gene toolkit for future gene expression studies in S. chinensis, particularly for the functional characterization of genes involved in lignan biosynthesis and abiotic stress responses. Full article

Objectives: This study investigated a variant, RPGR NM_001034853.2 c.1307G>A, p.[Gly436Asp, p?], in a large Irish pedigree with severe X-Linked Retinitis Pigmentosa (XLRP). The effect of the variant on RNA splicing was interrogated using in vitro functional analysis to provide evidence of disease causality. Methods: Three related individuals presenting with XLRP underwent target-capture sequencing, together with confirmatory Sanger sequencing and cascade analyses, to identify candidate variants. In silico investigations were undertaken using SpliceAI (version 1.3.1) and Alamut Visual software (version 2.13), among others. Functional analyses using in vitro midigene splice assays employing gateway expression vectors were undertaken. Variant and wildtype RNA were amplified by RT-PCR to investigate effects on splicing. RPGR c.1307G>A was subsequently reclassified using ACMG/AMP and ClinGen SVI recommendations. Results: Midigene investigation confirmed a cryptic acceptor site is being utilised together with the cryptic branchpoint motif to excise intron 10 and 90 bases of exon 11, leading to a frameshift and the creation of a premature stop codon. No functional RPGR transcript is predicted to remain. Given evidence of aberrant splicing, the variant classification was upgraded to pathogenic. Conclusions: RPGR c.1307G>A leads to creation of a cryptic branchpoint within an exon, resulting in protein truncation with deleterious effect(s). To the best of our knowledge, this is the first variant that leads to creation of a cryptic branchpoint within an exon associated with any IRD. The results illustrate the importance of investigating the functional consequences of both coding and non-coding variants with a predicted impact on splicing to understand their pathogenicity. Full article

Concussions exert a massive cost on our economic and healthcare systems. Many of the most commonly employed neurocognitive measures in concussion assessment have been shown to be psychometrically problematic. Additionally, norms are established from largely male populations. The present study investigates the use of a validated and reliable measure of concussion sequelae, the mobile half-version of the Connors Continuous Performance Test 3rd Edition (CCPT-3), on a representative population to study the influence of sex and age on normative values collected at baseline. Baseline data were analyzed from 71,976 participants across a wide range of academic and athletic contexts, as well as healthcare settings. Multiple regressions examined the influence of sex as a function of age in different developmental groups: children, adolescents, young adults, adults, and older adults. Sex effects emerged during childhood, peaked during adolescence, and decreased in adulthood. Females showed better accuracy (fewer commission and omission errors), whereas males had faster response speeds (hit-rate RT). Effect sizes were generally in the small to very small range (sex effect sizes ranged from Cohen’s d = 0.02 to 0.39). The findings highlight the importance of accounting for sex and age in cognitive test performance and underscore the impact of correcting for even small effects when working with large samples. Full article

Cyhalofop-butyl, a widely used acetyl-CoA carboxylase (ACCase)-inhibiting herbicide, is increasingly resisted by Echinochloa crus-galli in paddy fields. However, the physiological and molecular mechanisms of this resistance have not been fully elucidated. In this study, a highly resistant population (HL-R, resistance index [RI] = 19.05) from Hulin and a susceptible population (HL-S, RI = 1.00) were used. The results showed that HL-R population exhibited broad-spectrum resistance to five herbicides (RI = 1.98–14.53). No target-site mutations were found in the ACCase gene; instead, its overexpression contributed to target-site resistance (TSR). Metabolic inhibitor assays confirmed non-target-site metabolic resistance (NTSR). Transcriptome and qRT-PCR analyses identified two detoxification-related differentially expressed genes (DEGs; CH01.636, BH02.4557) as key regulators. The activity of the antioxidant enzyme peroxidase (POD) was also associated with resistance. Collectively, the high resistance of HL-R to cyhalofop-butyl is synergistically driven by ACCase overexpression, POD activity and P450/GST-mediated metabolic detoxification. These findings provide theoretical guidance for the management of resistant barnyard grass. Full article

The genetic and antigenic diversity of H5Nx HPAI Gs/GD lineage continues to be a great challenge facing conventional inactivated vaccines. To overcome this challenge, a recombinant herpes virus of turkey (rHVT) vaccine expressing the viral protein 2 (VP2) of infectious bursal disease (IBD) and H5, rHVT+IBD+H5, was developed using computationally optimized broadly reactive antigen (COBRA) technology. In the current study, the protective efficacy of a commercially available vector trivalent vaccine rHVT+IBD+H5 using COBRA technology was assessed. A total of 120 commercial broilers were divided equally into six groups (G1B–G6B). The chickens in G1B–G3B were challenged with the most recent circulating HPAI-H5N1 clade 2.3.4.4.b Egyptian isolate (GenBank accession No. OQ933425) at 28 days old (DO), while the chickens in G4B and G5B were kept as vaccinated (as G1B and G2B, respectively) and non-challenged, and G6B was the non-vaccinated non-challenged group. In G1B, the chickens were vaccinated with Vaxxitek^® rHVT+IBD+H5 at 1 DO and boostered with a commercially available subunit Baculovirus bivalent inactivated H5+ND (Volvac^® B.E.S.T AI+ND) at 10 DO and had a 100% survival rate. The standalone vaccinated chicken G2B, using rHVT+IBD+H5 at 1 DO, had a highly significant survival rate (90%) vs. 0% (100% mortality) in the non-vaccinated challenged control, G3B. All the vaccinated groups had higher seroconversion at 45 DO especially using H5-coated antigen plates for the enzyme-linked immunosorbent assay (ELISA) test. The viral shedding titers and time were evaluated using a quantitative real-time polymerase chain reaction (RT-qPCR) in the collected oropharyngeal and cloacal swabs at 3, 5, 7, and 10 days post-challenge (DPC). In conclusion, vaccination with rHVT+IBD+H5 either as a standalone or when boostered with subunit Baculovirus bivalent inactivated ND+H5 resulted in 90 and 100% protection, respectively, without significant difference in the quantity and duration of viral shedding between both groups against HPAI-H5N1 clade 2.3.4.4.b experimental challenge in broilers. Full article

Rapid detection of foodborne pathogenic and spoilage microorganisms is critical for ensuring food safety and quality in liquid matrices. While Raman tweezers spectroscopy (RTS) enables label-free single-cell analysis, its application in high-throughput inline inspection faces a fundamental bottleneck: high flow rates required for efficiency induce severe motion blur and low signal-to-noise ratios (SNR), which blind automated control systems and destabilize optical trapping. To overcome this, we present a Spatiotemporal Video-Enhanced Raman Tweezers (SVERT) system integrating a deceleration-optimized microfluidic chip with a deep learning-based visual feedback loop. We propose a Local–Global Unified Denoising Network (LGU-Net) tailored to recover high-fidelity bacterial structures from low-SNR video streams, achieving a deterministic processing latency of ~0.49 ms. Experimental results demonstrate that SVERT improves the optical trapping success rate from 21.27% ± 2% to 91.47% ± 1.8% compared to raw video input, enabling a four-fold increase in spectral acquisition efficiency. Leveraging the acquired high-quality dataset, we achieved a classification accuracy of 96.74% across four bacterial species of relevance to food safety and quality. Crucially, we validated the system’s practical robustness by successfully isolating and tracking trace E. coli in an unpurified commercial beverage. This capability to effectively mitigate natural background interference demonstrates the system’s promising potential to be expanded for broader applications in liquid food safety screening. Full article

Mo–Re alloys serve as critical structural components for high-temperature nuclear reactors, and their irradiation degradation is closely related to the evolution of vacancy-type defects. In this study, heavy-ion and He-ion irradiations were performed under RT to introduce an average displacement damage of 3.5 dpa within the 1 μm-thick surface layer of Mo–Re alloys with Re content up to 47 wt.%. PALS, SPB-DBS and CDB techniques were employed to characterize the size, concentration, depth distribution and local chemical environment of irradiation-induced vacancy-type defects. The results demonstrate that the longer lifetime component of irradiated Mo–Re alloys ranged from 262 to 280 ps, corresponding to medium-sized vacancy clusters. The S parameter of all specimens increased significantly from approximately 0.42 to 0.50, with negligible differences (<0.01) among various Mo–Re alloys. No distinct characteristic peak of Re was observed near 17 × 10⁻³ m₀c at the vacancy sites, which was inconsistent with simulation predictions. Mo–Re alloys exhibit similar vacancy-type defect features to pure Mo, implying weak interactions between Re solute atoms and vacancy-type defects under RT irradiation. Full article

TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) are plant-specific regulators involved in growth, development, and responses to abiotic stresses, yet their roles in halophytes remain largely unexplored. In this study, we performed a genome-wide identification of TCP family members in the extreme halophyte Salicornia europaea, uncovering 15 non-redundant genes (SeurTCPs) classified into PCF, CIN, and CYC/TB1 subfamilies. Gene structure and conserved motif analyses revealed that SeurTCPs are largely intronless and maintain the canonical TCP domain, while showing subfamily-specific variations in motif composition and secondary/tertiary structures. Promoter analysis identified abundant stress and hormone-responsive cis-elements, particularly ABRE and STRE, suggesting potential involvement in salt stress signaling. Protein–protein interaction network prediction highlighted CIN and PCF members as hub nodes, indicating central roles in growth and stress response regulation. Quantitative Real-Time Reverse Transcription Polymerase Chain Reaction (qRT-PCR) analysis showed that most SeurTCP genes were responsive to salinity treatment, although the extent of transcriptional variation differed among subfamilies. Collectively, our results indicate that SeurTCPs balance conserved structural functions with subfamily-specific regulatory roles, contributing to S. europaea adaptation to extreme saline environments. This study provides valuable candidate genes for elucidating plant salt tolerance mechanisms and for potential crop improvement. Full article

Introduction: The temperature of rivers in the Iberian Peninsula has increased due to global warming. In addition, these rivers are polluted by contaminants of emerging concern, such as polycyclic aromatic hydrocarbons (PAHs). Higher temperatures and pollution concurrently impose threats to the Iberian Peninsula’s endemic species, including the brown trout (Salmo trutta), a cold-water species widely used in ecotoxicological studies. Because the liver is the main biotransformation organ, and is particularly sensitive to both chemical and temperature changes, in vitro liver models may represent valuable alternatives for assessing combined stressor effects, complying with the 3Rs principle. Objective: In line with the above, the present study aimed to evaluate the combined effects of a 4 °C temperature increase and the model PAH benzo[k]fluoranthene (B[k]F) on fish liver cells using a primary brown trout hepatocyte culture as a model. Methodology: Primary hepatocytes were seeded in 6-well plates at a density of 1.0 × 10⁶ cells/mL and exposed for 48 h to 1, 10, and 20 µM B[k]F at 18 °C (normothermia) and 22 °C (warming scenario). Cell viability was assessed using trypan blue, alamarBlue, and lactate dehydrogenase (LDH) assays. Cytochrome P450 (CYP)1A was evaluated in terms of its gene expression by RT-qPCR and its protein expression through immunocytochemistry (ICC). The immunostaining was quantified using a score system which considered five intensity staining levels. Results: Exposure to B[k]F and to the higher temperature increased LDH leakage without interaction effects. In contrast, the other viability assays did not show significant differences across conditions. Regarding CYP1A, both gene and protein expression increased with all B[k]F concentrations in relation to the controls, but were not influenced by temperature. Notably, the lowest B[k]F concentration (1 µM) elicited the highest CYP1A gene expression, suggesting a non-monotonic response. Conclusions: Overall, the model was responsive to both temperature (4 °C) increase and to B[k]F, validating its usefulness for assessing liver pollutant effects in the context of global warming. These findings support the application of fish primary hepatocyte models as relevant tools in ecotoxicology under environmentally realistic multi-stressor scenarios. Full article

While the cerebellar dopaminergic system is suggested to be implicated in neurodevelopmental disorders, especially autism spectrum disorder (ASD), the details of its disturbances remain unclear. We performed a comparative analysis of human (GTEx) and mouse (GSE144046, GSE144277) transcriptomes, complemented by RT-qPCR in DAT-KO rats, to identify dopaminergic gene associations in the normal cerebellum and neurodevelopmental disorder models. Pairwise dopaminergic gene correlations were generally weak, with a slight increase in interaction complexity in ASD models. However, weighted gene co-expression network analysis identified a robust gene module involving Comt, which was consistently associated with synaptic translation across mouse datasets. These associations reflect regulatory processes in the whole cerebellum, which is commonly represented in rodent studies but absent in human data, which are acquired in studies of cerebellar subregions. ASD modeling exerted contrasting effects: Cul3 haploinsufficiency increased the number of genes involved in the module with a decrease in connectivity, while Mbd5 haploinsufficiency led to module collapse. These findings confirm neurodevelopmental disorders as a heterogeneous condition where divergent backgrounds uniquely rewire cerebellar dopaminergic networks. Considering the cerebellum’s role in ASD and that some ASD medications target the dopamine system, further investigation of these identified trends may support the development of more personalized therapeutic approaches. Full article

Monolayer and few-layer MoS₂ are promising two-dimensional electronic materials, but proton irradiation can trigger ultrafast electronic excitation and charge transfer before defect formation. Here, real-time time-dependent density functional theory (RT-TDDFT) is used to investigate proton-induced electronic stopping and localized charge capture in monolayer and bilayer MoS₂ under normal incidence. Four impact positions are examined in monolayer MoS₂, namely, the hollow channel, the Mo–S bond center, and two trajectories close to Mo and S atoms. Under hollow channel incidence, the stopping power shows a non-monotonic dependence on proton velocity. When comparing the different trajectories, the hollow channel path gives the lowest stopping power, whereas the Mo–S bond center path gives the highest values, indicating strong sensitivity to the in-plane valence charge distribution. By contrast, the time-averaged localized captured charge decreases with increasing velocity and is generally largest for the close to Mo trajectory. Under the same hollow channel condition, the monolayer stopping power exceeds the bilayer value in the main stopping region, whereas the bilayer generally shows slightly enhanced localized charge capture. These results show that electronic stopping and localized charge capture are distinct but coupled microscopic components of proton-induced electronic response in MoS₂ and provide first-principles insight relevant to ion-beam processing and radiation-tolerant two-dimensional devices. Full article

Kadsura heteroclita (Roxb.) Craib, commonly known as “Xuetong”, is a traditional Tujia ethnomedicine with anti-rheumatoid arthritis (RA) activity, and schizanlactone E (Xuetongsu) is its major bioactive component whose biosynthetic pathway remains uncharacterized. As a cycloartane-type tetracyclic triterpenoid, Xuetongsu’s biosynthesis is likely to involve multiple oxidation steps. Cytochrome P450 (CYP450) is a versatile monooxygenase encoded by a large and diverse gene superfamily and plays a critical role in various oxidation reactions in plants’ secondary metabolism. In this study, 367 KhCYP450s were identified and systematically analyzed for their physicochemical properties, phylogenetic analysis, conserved motifs, gene structures, collinearity, and cis-acting elements. Weighted gene co-expression network analysis (WGCNA) revealed a turquoise module strongly associated with Xuetong root tissue, which had the highest Xuetongsu accumulation; 32 candidate KhCYP450s within this module were screened via correlation analysis between gene expression and xuetongsu content and partially validated by qRT-PCR. Five of these candidates showed significant homology with known triterpenoid biosynthetic genes via protein structure analyses. This study deepened our comprehension of the CYP450 superfamily in Xuetong and provided a valuable reference for further research on the biosynthesis of Xuetongsu. Full article

Optical remote sensing image object detection is widely applied in industrial monitoring, infrastructure maintenance, and intelligent scheduling, yet its accuracy is limited by complex background interference, dense multi-scale object distributions, and loss of small object details. To address these challenges, this paper proposes the ORSSO-DETR model based on RT-DETR, which improves the hybrid encoder’s ability to adapt to the characteristics of optical remote sensing images. The proposed method integrates the C3K2 multi-scale fusion module with the FCM feature complementary mapping module to enhance feature fusion, introduces a dynamic convolution-based DynamicConv downsampling module and the EUCB upsampling technique to improve multi-scale feature retention and texture detail preservation, optimizes the encoder structure by preprocessing high-level features with 1 × 1 convolution, and replaces the original normalization layer in the Transformer with the Dynamic Tanh module to strengthen nonlinear expressive capability. Experimental results show that the proposed method achieves 96.2% mAP@50 and 64.4% mAP@50:95 on the NWPU VHR-10 dataset, improving by 3.9% and 2.9% over the baseline, respectively. On the RSOD dataset, it achieves 97.6% mAP@50 and 71.2% mAP@50:95, with improvements of 2.5% and 2.8%, respectively. These results validate the effectiveness of the proposed method in improving small object detection accuracy in optical remote sensing images. Full article

The rhizosphere microbiome and phytohormone signaling are critical determinants of plant growth and stress resilience. This study evaluated the combined effects of Streptomyces sp. HU2014 and coronatine (COR) on maize (Zea mays L.) seedlings. Four treatments were established: control (CK), COR seed soaking (Cor), HU2014 soil inoculation (S), and combined S + Cor (SCor). Growth parameters, chlorophyll content, and antioxidant/oxidative stress markers were measured, and root and leaf transcriptomes, together with root metabolomes, were compared between SCor and CK, followed by qRT-PCR validation. Compared with CK, SCor treatment significantly increased stem diameter (~60%), plant height (~20%), and relative chlorophyll content (SPAD, ~50%). Soluble sugar levels were elevated by over 40% in both leaves and roots, accompanied by tissue-specific modulation of antioxidant enzymes. Transcriptomic analysis of SCor vs. CK revealed 2459 differentially expressed genes (DEGs) in leaves and 3444 DEGs in roots; leaves exhibited upregulation of photosynthetic pigment metabolism (porphyrin and carotenoid pathways) and volatile defense compounds (alkaloids and monoterpenoids), whereas roots showed enrichment in phenylpropanoid/flavonoid biosynthesis, benzoxazinoid synthesis, and starch/sucrose metabolism. Metabolomics of SCor vs. CK identified 526 differentially accumulated metabolites (DAMs) in roots, with significant enrichment in aminoacyl-tRNA biosynthesis, phenylalanine metabolism, and linoleic acid metabolism. Integrative multi-omics analysis further revealed that the JA precursor 13-epi-12-oxo-phytodienoic acid co-clustered with stress-responsive transcription factors (e.g., DREB1C), while tricarboxylic acid (TCA) intermediates and phenylpropanoid metabolites were linked to energy and lignin biosynthesis genes. qRT-PCR confirmed the expression trends of 14 out of 15 tested genes. Collectively, combined HU2014 and COR application triggers tissue-specific transcriptional and metabolic reprogramming in maize, coupling JA-mediated stress signaling with enhanced carbon metabolism and secondary defense compound synthesis to promote rhizosphere adaptation and seedling vigor. Full article

Object detection in remote sensing imagery plays an irreplaceable role in critical fields such as military reconnaissance and disaster monitoring. However, when dealing with minute targets characterised by an extremely low pixel proportion, a lack of textural information, and severe background interference, existing algorithms still face the challenge of balancing detection accuracy with computational efficiency. To address this, this paper proposes a lightweight frequency-domain-aware end-to-end detection model, LE-DETR, based on an improved version of RT-DETR. Firstly, a Lightweight Feature Extraction Module (LFEM) is designed. Through a heterogeneous dual-path architecture and reparameterisation techniques, it significantly reduces computational complexity whilst enhancing the capture of fine-grained spatial features. Secondly, an Efficient Spatio-Frequency Fusion Module (ESFFM) is introduced. This utilises a multi-head self-attention mechanism to construct a global view whilst combining the Fourier transform to reconstruct target features from a frequency-domain perspective, thereby effectively suppressing background noise and enhancing the target’s edge signals. Finally, we propose the Efficient Frequency-Aware Fusion Feature Pyramid Network (EFAM-FPN), which utilises SPD Conv to mitigate the loss of key features during downsampling and introduces a frequency-domain attention mechanism to suppress complex background noise, thereby improving the model’s detection accuracy for extremely small objects. The experimental results show that, whilst reducing the number of parameters by 41.7% compared to the baseline model, LE-DETR achieved improvements of 2.6%, 1.7% and 2.4%, respectively, in the mAP50 metric across the three mainstream remote sensing datasets—VisDrone2019, NWPU VHR-10 and DIOR. This demonstrates an effective balance between detection accuracy and inference efficiency, fully validating its robustness and practical value in complex remote sensing application scenarios. Full article