Search Results (675)

Background: Nanopore sequencing produces ionic current signals that are sensitive to chemical modifications in DNA and RNA molecules. However, accurate modification detection remains challenging due to limited labeled data and variability across experimental conditions. Methods: We present a scalable unsupervised framework for modification discovery that learns reference signal distributions from unmodified sequences using a CNN–Transformer variational autoencoder (VAE). The model is trained on large-scale data via streaming sampling and k-mer-aware soft balancing to ensure robust signal representation. At inference, candidate nucleotides are scored using the VAE reconstruction error, and read-level signals are aggregated to produce site-level modification evidence. Results: On controlled DNA oligonucleotide datasets, models trained on unmodified sequences achieve strong discrimination when evaluated on modified oligos. In contrast, performance decreases in cell line samples when models trained on unmodified whole-genome-amplified (WGA) DNA and in vitro-transcribed (IVT) RNA are evaluated on natively modified (5mC/m6A) data, reflecting the impacts of biological noise and heterogeneity. Despite reduced classification accuracy, site-level anomaly score profiles exhibit peak-like patterns that correspond to known modification-enriched regions. Conclusions: These findings demonstrate the feasibility of large-scale unsupervised reference modeling for de novo modification detection, while underscoring the challenges in translating models built from synthetic oligo datasets into robust genome-wide modification detection. Full article

The effective control of foodborne salmonellosis relies on the rapid and reliable detection and identification of the pathogen. Reliable detection tools for identifying the most common Salmonella serovars should translate to a considerable alleviation of the health burden attributed to Salmonella. We have developed a PCR assay for the rapid identification of colonies of Salmonella enterica serovar Thompson, a common serovar. Genomic analyses of publicly available sequences of Salmonella Thompson revealed the presence of a unique, Thompson-specific fragment, which we have used to design a pair of oligonucleotides, STho-F and STho-R, for the PCR amplification of an 808 bp DNA fragment. Using crude DNA extracts, the 808 bp fragment was detected in 77 out of 78 isolates of S. Thompson (sensitivity = 98.7% n = 78 isolates) but not in any of the non-Salmonella organisms tested (n = 100; 100% specificity) nor in non-Thompson Salmonella serovars (n = 100; 100% specificity). The sensitivity (inclusivity) and specificity (exclusivity) indices of the PCR assay for S. Thompson met the standard regulatory requirements. The Thompson primer pair was compatible with other primers pairs in a multiplex PCR designed for three other common Salmonella serovars. Colonies belonging to the Enteritidis serovar (n = 100), Heidelberg serovar (n = 100), Typhimurium serovar (n = 100), and Thompson serovar (n = 77) were correctly designated, indicating excellent inclusivity and exclusivity scores for all four Salmonella serovars tested in a single multiplex PCR. Full article

A quartz crystal microbalance-based biosensor for the specific detection of the first transgenic common bean (L.) cultivar (BRS FC401 RMD) with resistance to Bean golden mosaic virus (BGMV) was developed. The immobilization chemistry relies on the strong bond between the thiolated probe and the gold electrode surface. The probe sequence is internal to a region of the BGMV rep gene that was introduced into the common bean genome. The sensor’s analytical performance was determined using synthetic oligonucleotides. Real samples of transgenic and wild-type bean seeds were also tested. Sample pretreatment consisted only of enzymatic fragmentation, followed by a thermal denaturation step combined with blocking oligonucleotides. Different biosensor regeneration approaches were studied. Immobilization showed good reproducibility (CV% of 5.8%). The biosensor proved specific for both synthetic oligonucleotides and non-amplified genomic DNA. A linear detection range of 0–1.4 ng/µL was observed, with a detection limit of 0.18 ng/µL. Three sequential detections were performed without loss of surface activity. The results demonstrate the biosensor’s potential for direct, real-time, label-free detection of DNA samples for field screening of transgenic common bean cultivars. Full article

SPR (surface plasmon resonance) biosensor–based analytical methods enable rapid, straightforward, and cost-effective detection of DNA oligonucleotides. However, the detection limits of currently available SPR biosensors for BCR–ABL gene oligonucleotides remain too high to reliably detect sub-nanomolar concentrations. This study presents a new signal-enhancement approach for SPR DNA biosensors based on a gold nanoparticle (AuNP) sandwich assay. In this work, we demonstrated that AuNP-modified oligonucleotides can serve as labels that significantly amplify the SPR biosensor response in a sandwich-type SPR DNA biosensor. The analytical characteristics of the developed AuNP-labeled biosensor for detection of BCR–ABL fusion gene oligonucleotides were studied. The AuNP-labeled biosensor exhibited a detection limit of 80 pM, which is significantly lower than that of a traditional label-free SPR biosensor (50 nM). The measurement error for BCR–ABL target detection was significantly lower with the AuNP-labeled biosensor than with the label-free SPR biosensor. The conditions of synthesis of AuNPs by citrate reduction of AuCl₃ that allow the monodisperse size distribution and absence of AuNP aggregation were established as well. Based on the obtained data, we conclude that a sandwich assay employing AuNP-modified oligonucleotides as labels is a promising approach for the highly sensitive detection of genetic markers. The developed AuNP-labeled DNA biosensing approach can be adapted to enhance the signal in other DNA hybridization-based SPR biosensors. Full article

Duchenne muscular dystrophy (DMD) is a severe X-linked neuromuscular disease (NMD) caused by loss-of-function mutations in the DMD gene. RNA-based therapies, especially antisense oligonucleotides (ASO)-mediated exon skipping and adenosine deaminase acting on RNA (ADAR)-guided RNA editing, have emerged as complementary approaches that modulate pre-mRNA splicing or correct transcripts without altering genomic DNA. Current phosphorodiamidate morpholino oligomer (PMO) drugs targeting exons 51, 53, and 45 provide mutation-class-specific benefit. At the same time, next-generation delivery strategies (e.g., peptide-conjugated PMOs (PPMOs), antibody–oligonucleotide conjugates (AOC), and endosomal-escape vehicles) aim to improve skeletal, cardiac, and diaphragm exposure. In parallel, RNA editing strategies offer a route to correct select nonsense or missense variants at the base level and may, in principle, restore near-native dystrophin expression. Meaningful translation of these modalities requires predictive large-animal models. A genome-edited microminipig (MMP) bearing DMD exon-23 mutations faithfully recapitulates hallmark features of human DMD. That includes early locomotor deficits, elevated serum creatine kinase (CK) and cardiac troponin T, progressive myocardial fibrosis, and a decline in left-ventricular ejection fraction (LVEF), while maintaining a manageable lifespan of approximately 30 months suitable for long-term studies. In particular, the MMP model provides a practical platform for addressing the persistent challenge of efficient therapeutic delivery to the heart and diaphragm through longitudinal dosing, imaging, and biopsy. In this review, we synthesize clinical progress in exon skipping, outline the promise of RNA editing, and integrate recent insights from Duchenne muscular dystrophy model for microminipigs (DMD-MMPs) as an advanced surrogate for preclinical development and translational evaluation. Full article

Large language models (LLMs) hold promise for automated extraction of structured biological information from scientific literature, yet their reliability in some domain-specific tasks, such as DNA probe parsing remains underexplored. We developed a verification-focused, schema-guided extraction pipeline that transforms unstructured texts from scientific articles into a normalized database of oligonucleotide probes, primers, and associated metadata. The system combined multi-turn JSON generation, strict schema validation, sequence-specific rule checks, and a post-processing recovery module that rescues systematically corrupted nucleotide outputs. Benchmarking across nine contemporary LLMs revealed distinct accuracy–hallucination trade-offs, with the context-optimized Qwen3 model achieving the highest overall extraction efficiency while maintaining low hallucination rates. Iterative prompting substantially improved fidelity but introduced notable latency and variance. Across all models, stable error profiles and the success of the recovery module indicated that most extraction failures stem from systematic and correctable formatting issues rather than semantic misunderstandings. These findings highlight both the potential and the current limitations of LLMs for structured scientific data extraction. The research provides a reproducible benchmark and extensible framework for future large-scale curation of molecular biology datasets. Full article

Aptamers, short single-stranded DNA or RNA oligonucleotides, are emerging as transformative tools in cardiology for the diagnosis, treatment, and theranostics of cardiovascular diseases (CVDs). This review highlights their dual utility. In diagnostics, aptamers enable the construction of highly sensitive biosensors for key cardiac biomarkers (e.g., troponins, myoglobin, C-reactive protein, natriuretic peptides), outperforming conventional assays and enabling early detection and point-of-care testing. Therapeutically, aptamers offer targeted, controllable, and reversible anticoagulation, as demonstrated by clinical-stage candidates like BT200 (anti-vWF) and NU172 (anti-thrombin), whose action can be rapidly reversed with antidote oligonucleotides. Furthermore, aptamers serve as precision delivery vehicles (e.g., Gint4.T, RNA-Apt30) for transporting therapeutic peptides or nucleic acids specifically to cardiomyocytes. Recent integration with nanomaterials (quantum dots, graphene, liposomes, DNA origami) has led to advanced biosensing and drug delivery platforms. Despite challenges like stability and the polyethylene glycol (PEG) immunogenicity, ongoing clinical trials underscore the significant potential of aptamer technology to bridge precise diagnostics and targeted therapy, paving the way for innovative, personalized CVD interventions.) Full article

This study developed and validated a Self-competitive Fishing (SCF) primer qPCR system as a rapid, cost-effective alternative to next-generation sequencing (NGS) for detecting POLE exonuclease domain mutations (EDMs) in endometrial cancer. The system detects 11 pathogenic POLE EDMs using SuperSelective primers combined with wild-type-blocking oligonucleotides that prevent amplification of wild-type DNA, thereby enhancing mutant DNA detection. The validation process involved comparing specificity using genomic DNA from tumors with known POLE mutations identified by NGS. Sensitivity testing used POLE-mutated DNA diluted in wild-type DNA, while precision was confirmed by analyzing 86 endometrial cancer samples against NGS results. The SCF qPCR system demonstrated superior specificity compared to the original SuperSelective primer-based qPCR, achieving 1% mutation-detection sensitivity across various mutation points. Importantly, results from all endometrial cancer cases showed complete concordance with NGS analysis for the 11 pathogenic POLE-EDM points tested. This cost-effective and efficient SCF primer qPCR system provides an accessible method for routine molecular classification of endometrial cancer in clinical settings, offering a practical alternative to NGS for detecting pathogenic POLE mutations and supporting clinical decision-making. Full article

Recent advances in molecular genetics, nucleic acid synthesis, and bioinformatics have provided novel opportunities for plants’ protection against insect pests. Currently, both DNA and RNA serve as active insecticidal ingredients, transcending their traditional role as carriers of genetic information. This novel activity is achieved through two fundamentally distinct mechanisms. The first one is DNA containment (DNAc), employing oligonucleotide insecticides based on contact unmodified antisense DNA biotechnology (CUADb), also known as ’genetic zipper’ technology. The second one is RNA interference (RNAi), employing RNA biocontrols based on double-stranded RNA (dsRNA) technology. The investigation of the molecular mechanism underlying the antisense activity of nucleic acids emerged in the early 1960s. While the antisense effects of RNA in gene silencing through interference (RNAi) was documented in the late 1990s as antiviral immune responses in nematodes, the CUADb antisense approach initially emerged as a powerful strategy for pest control against lepidopterans in 2008. The CUADb approach relies on disrupting rRNA biogenesis and ribosome production, while RNAi shows the best results in mRNA degradation and no efficient result is known for rRNA. The efficacy of these approaches appears to be species dependent. For example, CUADb demonstrates optimal activity against Sternorrhyncha (e.g., aphids, mealybugs, psyllids, and scale insects), thrips, and mites. In turn, the RNAi strategy shows a strong insecticidal potential against beetles from the Tenebrionidae and Chrysomelidae families. Here, we will review the differences between the two technologies, their mechanisms of action and the current challenges facing their adoption. Full article

Background/Objectives: Melanoma is the deadliest form of skin cancer. Resistance to alkylating agents such as Temozolomide (TMZ) and Dacarbazine (DTIC) limits their clinical benefit, as these drugs remain palliative options when immunotherapies and targeted treatments fail. CSA/ERCC8 is a key component of transcription-coupled nucleotide excision repair (TC-NER), a pathway responsible for removing UV-induced DNA lesions. In principle, loss of a DNA repair factor would be expected to increase carcinogenesis. However, although CSA loss-of-function causes Cockayne Syndrome (CS), affected patients do not exhibit increased skin cancer incidence, suggesting that CSA impairment promotes apoptosis rather than tumor development. This paradox raises the possibility that CSA inhibition may selectively target melanoma cell survival pathways. Methods: The expression of CSA/ERCC8 was analyzed by qRT-PCR and Western blot. ERCC8 was silenced using antisense oligonucleotides. Cell viability, apoptosis, cell cycle progression, drug sensitivity, and DNA damage were assessed by functional assays, including IC50 determination and Bliss analysis for drug interactions. Results: We identified CSA/ERCC8 as a driver of melanoma chemoresistance. CSA was markedly overexpressed in primary and metastatic melanoma cells. ERCC8 silencing reduced proliferation, induced apoptosis, and significantly enhanced sensitivity to low doses of TMZ and DTIC while sparing normal cells. Conclusions: CSA represents a promising therapeutic target to overcome chemoresistance in melanoma. Its inhibition enhances the efficacy and selectivity of alkylating agents, supporting its potential as a salvage strategy for refractory disease and warranting further preclinical and clinical investigation. Full article

Understanding interactions between cations and DNA is essential for elucidating the structural dynamics of this fundamental biomolecule. While B-DNA is well known to dominate in long genomic DNA under physiological ionic conditions, its stability in very short DNA fragments—particularly in dilute solutions and in crude oligonucleotide preparations—has remained largely unexplored. Previous spectroscopic studies have primarily focused on long DNA, highly purified oligonucleotides, or high-salt environments, where collective polyion effects dominate. In contrast, the present results demonstrate that even in the absence of chain overlap and under low-salt conditions, Mg²⁺ ions efficiently stabilize the B-form by screening phosphate–phosphate electrostatic repulsion at the intrachain level. The ability to induce an A-to-B transition in crude, ultra-short DNA fragments highlights the fundamental role of divalent counterions in governing DNA conformation and establishes a lower bound for the length scale at which B-DNA can be stabilized. These findings are particularly relevant for dilute biological systems, fragmented DNA samples, and analytical protocols where short DNA fragments and low ionic strength are unavoidable. Full article

Epilepsy is a heterogeneous neurological disorder with a strong genetic basis, yet recent evidence underscores the critical role of epigenetic mechanisms in its pathogenesis. This review synthesizes current knowledge on how chromatin remodeling, histone modifications, DNA methylation, and transcriptional regulation intersect with classical channelopathies and signaling pathways. We emphasize how epigenetic dysregulation contributes to neuronal excitability and network plasticity, particularly through interactions with mTOR, PI3K-AKT, and GABAergic signaling cascades. The convergence of genetic mutations and epigenetic modifications creates a dynamic landscape in which environmental factors can modify gene expression and contribute to the development of epilepsy. Emerging therapeutic strategies—including epigenetic drugs (HDAC inhibitors, DNMT inhibitors), CRISPR/dCas9-based epigenome editing, and multi-omics approaches—offer promising avenues for precision medicine. This review provides a comprehensive synthesis of genetic and epigenetic mechanisms in epilepsy, examining how these layers interact to produce disease phenotypes and discussing the therapeutic implications of this multilayered regulation. Full article

Telomere length is a crucial marker of cellular aging and genomic stability, with significant implications for age-related diseases and cancers. This study introduces an improved quantitative PCR (qPCR) method for measuring absolute telomere length, addressing the need for accurate and high-throughput assessment in both clinical and research settings. Novel primers were designed for the single-copy gene interferon beta (IFNB1) to serve as an internal control, alongside a series of single-stranded oligonucleotide standards to establish a calibration curve. This approach allows for precise quantification of telomere length in kilobases per single copy gene copy number per chromosome. We validated this method using DNA samples from peripheral blood and buccal swabs from 17 healthy human volunteers, as well as umbilical cord blood from 9 healthy newborn babies, demonstrating its high linearity and reproducibility. Our findings indicate that this improved qPCR technique provides a rapid, cost-effective, and accurate means of measuring absolute telomere length, thereby facilitating large-scale studies and enhancing clinical diagnostics related to telomere biology. Full article

Background/Objectives: Interleukin-17A (IL-17A) is a key pathogenic cytokine in autoimmune arthropathies. Current monoclonal antibody inhibitors targeting the IL-17/IL-17RA axis demonstrate clinical efficacy but face significant limitations, including immunogenicity, the loss of therapeutic response, and cold-chain storage. Our study evaluated oligonucleotide aptamers targeting IL-17A and its receptor as an alternative to monoclonal antibodies to suppress an IL-17A-induced inflammatory response in cell models relevant to immunoinflammatory rheumatic diseases. Methods: We examined three aptamers: 2′-F-RNA aptamers Apt21-2 and Apt3-4 specific to IL-17A and DNA aptamer RA10-6 targeting the receptor of IL-17A. Their ability to suppress IL-17A functional activity was assessed in peripheral blood mononuclear cells (PBMCs) from healthy donors and personalized fibroblast-like synoviocytes (FLSs) from patients with axial spondyloarthritis (axSpA) and rheumatoid arthritis (RA). Inhibition was measured by quantifying IL-6 and MMP-13 secretion using ELISA and flow cytometry, using secukinumab as a reference control. Results: In PBMC, all aptamers suppressed IL-17A-stimulated IL-6 secretion and cell proliferation in a concentration-dependent manner (17–200 nM), with a 65–85% efficacy, comparable to that of secukinumab. In axSpA-derived FLS, we observed time-dependent efficacy: At 4 h, all three aptamers suppressed IL-6 to the same extent as secukinumab; at 24 h, RA10-6 maintained high efficacy while Apt21-2 and Apt3-4 showed reduced activity. A combination of receptor-targeting RA10-6 with anti-IL-17A aptamers resulted in synergistic IL-6 suppression. All aptamers reduced MMP-13 to basal levels. RA-derived FLS showed diminished responses to all inhibitors. Conclusions: Aptamers demonstrate high specificity and sustained efficacy in suppressing IL-17A signaling for an in vitro model of spondyloarthritis, with superior performance over antibodies. Disease-dependent differential efficacy in RA FLS reflects heterogeneity consistent with limited clinical anti-IL-17 efficacy in RA. These findings show the strong potential of the studied aptamers as an alternative to monoclonal antibodies for IL-17-associated inflammatory arthropathies, particularly spondyloarthritis. Full article

Hepatocellular carcinoma (HCC) represents the second leading cause of cancer mortality worldwide and is mostly caused by hepatitis B virus (HBV) infection. HBV can induce HCC by an indirect mechanism of continuous necro-inflammation, contributing to hepatocyte damage and promoting cancer, as well as by viral intrinsic factors. Among them, the major contributors to the development of HBV-related HCC are represented by (i) HBV DNA integration in genes modulating cell proliferation, (ii) HBV pro-oncogenic proteins, such as HBx and HBs, and (iii) the accumulation of viral mutations, enhancing the tumorigenic features of HBV proteins. The currently available antiviral treatments, based on the usage of Nucleos(t)ide analogs (NUCs), substantially control HBV replication. However, even a successful NUC treatment does not completely abrogate HCC risk, since it rarely allows achievement of an HBV functional cure, the therapeutic end-point associated with HBsAg loss and more favorable liver outcomes. To date, novel therapeutic strategies based on innovative direct antivirals (nucleic acid polymers, small interfering RNAs, antisense oligonucleotides, covalently closed circular DNA (cccDNA) inhibitors, and capsid assembly modulators) and immune-therapeutics (therapeutic vaccines, checkpoint inhibitors, and Toll-like receptor agonists) are under evaluation in clinical trials. These approaches are showing promising data in terms of an HBV functional cure, thus representing novel strategies that could be beneficial for reducing the burden of HBV-related HCC. Lastly, further efforts in drug development are necessary to identify new compounds that could achieve a sterilizing HBV cure, implying the complete elimination of cccDNA and integrated HBV DNA, the only end-point that completely eradicates HBV and its related oncogenic risk. Full article