SENSORS AND ACTUATORS B-CHEMICAL
·
2024
·
IF 8
Xin Gao,Yun Yin,Junyu Xie,Shaohua Gong,Xuemei Li
CRISPR/Cas13a system has been combined with different isothermal amplification strategies for RNA analysis. Nevertheless, expensive enzymes are usually indispensable in these isothermal amplification methods, which increases the detection cost. In this study, a CRISPR/Cas13a-mediated triple signal amplification strategy has been designed for selective, sensitive and inexpensive detection of viral RNA. SARS-CoV-2 S RNA activated CRISPR/Cas13a to cut hairpin DNA, triggering catalytic hairpin assembly (CHA) and subsequent hybridization chain reaction (HCR) to form a long-stranded Y-type DNA. G-quadruplexes were distributed on the branch of the long-stranded DNA and bound with N-methyl mesoporphyrin IX (NMM) to enhance the fluorescence signal. SARS-CoV-2 S RNA was specifically detected with a limit of detection (LOD) of 285 fM. Notably, no additional expensive enzymes were required in the entire detection process. The CRISPR/Cas13a-mediated triple signal amplification strategy holds great potential for specific and sensitive viral RNA detection with low cost.
研究领域:
生物技术
·
分子生物学
·
病毒学
✓ 已验证
产品关联
DOI
高影响因子
Cancer Communications
·
2024
·
IF 20.1
Wei Mu,Pengfei Gu,Huating Li,Jinjin Zhou,Yulun Jian,Weiping Jia,Yang Ge
Background Benzo[a]pyrene (B[a]P), a carcinogen pollutant produced by combustion processes, is present in the western diet with grilled meats. Chronic exposure of B[a]P in hepatocellular carcinoma (HCC) cells promotes metastasis rather than primary proliferation, implying an unknown mechanism of B[a]P-induced malignancy. Given that exosomes carry bioactive molecules to distant sites, we investigated whether and how exosomes mediate cancer-stroma communications for a toxicologically associated microenvironment. Method Exosomes were isolated from B[a]P stimulated BEL7404 HCC cells (7404-100Bap Exo) at an environmental relevant dose (100 nmol/L). Lung pre-education animal model was prepared via injection of exosomes and cytokines. The inflammatory genes of educated lungs were evaluated using quantitative reverse transcription PCR array. HCC LM3 cells transfected with firefly luciferase were next injected to monitor tumor burdens and organotropic metastasis. Profile of B[a]P-exposed exosomes were determined by ceRNA microarray. Interactions between circular RNA (circRNA) and microRNAs (miRNAs) were detected using RNA pull-down in target lung fibroblasts. Fluorescence in situ hybridization and RNA immunoprecipitation assay was used to evaluate the “on-off” interaction of circRNA-miRNA pairs. We further developed an adeno-associated virus inhalation model to examine mRNA expression specific in lung, thereby exploring the mRNA targets of B[a]P induced circRNA-miRNA cascade. Results Lung fibroblasts exert activation phenotypes, including focal adhesion and motility were altered by 7404-100Bap Exo. In the exosome-educated in vivo model, fibrosis factors and pro-inflammatory molecules of are up-regulated when injected with exosomes. Compared to non-exposed 7404 cells, circ_0011496 was up-regulated following B[a]P treatment and was mainly packaged into 7404-100Bap Exo. Exosomal circ_0011496 were delivered and competitively bound to miR-486-5p in recipient fibroblasts. The down-regulation of miR-486-5p converted fibroblast to cancer-associated fibroblast via regulating the downstream of Twinfilin-1 (TWF1) and matrix metalloproteinase-9 (MMP9) cascade. Additionally, increased TWF1, specifically in exosomal circ_0011496 educated lungs, could promote cancer-stroma crosstalk via activating vascular endothelial growth factor (VEGF). These modulated fibroblasts promoted endothelial cells angiogenesis and recruited primary HCC cells invasion, as a consequence of a pre-metastatic niche formation. Conclusion We demonstrated that B[a]P-induced tumor exosomes can deliver circ_0011496 to activate miR-486-5p/TWF1/MMP9 cascade in the lung fibroblasts, generating a feedback loop that promoted HCC metastasis.
研究领域:
肿瘤学
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分子生物学
·
细胞生物学
·
癌症生物学
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药理学
·
毒理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOELECTROCHEMISTRY
·
2021
·
IF 5.373
Ya Zhang,Ning Li,Wenhao Ma,Mei Yang,Changjun Hou,Xiaogang Luo,Danqun Huo
MicroRNA exhibits different levels of expression in cancer and can affect the transformation, metastasis, and carcinogenesis of the cancer cell. Herein, we developed a novel kind of electrochemical microRNA biosensor based on two-dimensional nanomaterial of antimonene nano-flakes (AMNFs) and carbon quantum dots (CQDs) which were used as substrating to cadmium ion (Cd 2+ ) for specific detection of breast cancer-relevant biomarker-microRNA-21. Compared to graphene, the first principle energetic calculation shows that the AMNFs have completely a stronger force interaction with single strand (ssRNA), due to the antimonene has a more delocalized 5 s/5p orbital. After the addition of complementary microRNA, due to the low adsorption affinity of double-stranded RNA (dsRNA) to antimonene, the hybridized target is easy to desorb from the antimonene interface, and the oxidation peak of metal ions is significantly reduced. Results showed the microRNA-21 concentration can be detected from 100 aM to 1 nM, the limit of detection as low as 21 aM toward microRNA-21, which is 3 times lower than those of the established microRNA biosensors. The unique combination of not be attempted before existing sensing material which has special adsorption properties represents an approach to the detection of breast cancer. And it provides a promising method for early diagnosis, monitoring, and staging of breast cancer.
研究领域:
生物化学
·
纳米技术
·
癌症研究
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2018
·
IF 5.667
Lei Wang,Qi Xia,Ruiyuan Liu,Jinqing Qu
A red fluorescent probe, PFB, is designed and synthesized, which presents several remarkable features including a large Stokes shift (85 nm), long emission wavelength, high photostability and good biocompatibility. PFB also displays the solvent-dependent effect, which is verified by the density functional theory (DFT) calculations. Additionally, PFB can act as a fluorescent probe for cellular imaging with high uptake efficiency and highly sensitive detection toward RNA under low optimum concentration (1 μM), which enables it to achieve cancer cell tracing in vitro and tumor growth monitoring in vivo . The results of real-time tracking experiment reveal that PFB can be traced in stained HepG-2 cells for 11 generations and monitor tumor growth for 18 days. This work highlights the potential application of PFB as a promising alternative to the commercial RNA fluorescent probes.
研究领域:
生物化学
·
分子生物学
·
医学影像学
·
肿瘤学
✓ 已验证
产品关联
DOI
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2025
·
IF 8.5
Fang-Fang Liu,Hai-Yan Ren,Feng Liu,Wen-Xuan Qi,Chao-Peng Liang,Bang-Xian Zhang,Xiang-Jun Rao
Insects mount robust immune responses to reduce the efficacy of biopesticides. Identification of critical immune targets for biocontrol strategies is thus essential. However, immune targets in the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae), a major pest of corn, remain largely unexplored. Here, we investigated the immune response of S. frugiperda to the entomopathogen Beauveria bassiana (Hypocreales: Cordycipitaceae). Time-series RNA sequencing revealed 863, 717, and 165 differentially expressed genes (DEGs) at 24, 48, and 60 h post-infection, respectively. GO and KEGG enrichment analyses revealed that DEGs were significantly enriched in immune-related terms and pathways. Among the DEGs, the gene encoding immulectin SfIML-1 , a C-type lectin with dual carbohydrate recognition domains, was significantly upregulated during the early and intermediate stages of infection. Genome-wide analysis identified nine IML genes, with SfIML-1 as the only one highly responsive to B. bassiana . Recombinant SfIML-1 binds pathogen-associated molecular patterns and pathogens, enhancing cellular responses. RNAi of SfIML-1 decreased survival after fungal infection, while injection of recombinant SfIML-1 increased survival. Suppression of the NF-κB and JAK-STAT pathways diminished SfIML-1 expression in both fat body and Sf9 cells. These results identify SfIML-1 as a key pattern recognition receptor and a promising target for developing novel biocontrol strategies.
研究领域:
昆虫学
·
分子生物学
·
生物防治
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2024
·
IF 6.7
Benyu He,Lingfeng Wang,Xinyu Jin,Xing Zhang,Rui Sha,Yong Liang,Yawei Wang,Wenjing Xie,Jianbo Shi,Hanyong Peng
The detection of virus RNA in wastewater has been established as a valuable method for monitoring Coronavirus disease 2019. Carbon nanomaterials hold potential application in separating virus RNA owing to their effective adsorption and extraction capabilities. However, carbon nanomaterials have limited separability under homogeneous aqueous conditions. Due to the stabilities in their nanostructure, it is a challenge to efficiently immobilize them onto magnetic beads for separation. Here, we develop a porous agarose layered magnetic graphene oxide (GO) nanocomposite that is prepared by agglutinating ferroferric oxide (Fe3O4) beads and GO with agarose into a cohesive whole. With an average porous size of approximately 500 nm, the porous structure enables the unhindered entry of virus RNA, facilitating its interaction with the surface of GO. Upon the application of a magnetic field, the nucleic acid can be separated from the solution within a few minutes, achieving adsorption efficiency and recovery rate exceeding 90% under optimized conditions. The adsorbed nucleic acid can then be preserved against complex sample matrix for 3 days, and quantitatively released for subsequent quantitative reverse transcription polymerase chain reaction (RT-qPCR) detection. The developed method was successfully utilized to analyze wastewater samples obtained from a wastewater treatment plant, detecting as few as 10 copies of RNA molecules per sample. The developed aMGO-RT-qPCR provides an efficient approach for monitoring viruses and will contribute to wastewater-based surveillance of community infections.
研究领域:
环境科学
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纳米技术
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病毒学
·
分析化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BMC CANCER
·
2021
·Vol. 21
·期 1
·1-14
·
IF 4.43
Shi Yue,Liu Chong
Background Breast cancer (BC) is one of the major malignancies worldwide. Circular ribonucleic acids (circRNAs) are a class of conserved ribonucleic acid (RNA) molecules that play important roles in various diseases. Recently, circRNAs have been suggested to have diagnostic value and may function as potential diagnostic biomarkers for BC. Previously, hsa_circ_0043278 was found to be downregulated in human BC. However, its role in human BC has not yet been identified.Methods The levels of hsa_circ_0043278 in BC cell lines were verified by quantitative reverse transcription–polymerase chain reaction (qRT–PCR). The overexpression vector and short hairpin RNA (shRNA) of hsa_circ_0043278 were transfected into MDA-MB-231 and MCF-7 cells, respectively. The effects of hsa_circ_0043278 on tumour cell growth, migration and invasion were measured by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT), colony formation, wound healing and Transwell assays in vitro. A xenograft experiment was conducted to validate the inhibitory effect of hsa_circ_0043278 on tumour growth. The interaction between hsa_circ_0043278 and miR-455-3p was confirmed by a dual-luciferase reporter assay. Mimics and inhibitors of miR-455-3p were designed to confirm the influence of hsa_circ_0043278 on the hsa_circ_0043278/miR-455-3p/etoposide-induced gene 24 (EI24) axis.Results Hsa_circ_0043278 was downregulated in BC cell lines. Furthermore, overexpression of hsa_circ_0043278 notably decreased BC cell viability and inhibited BC cell migration and invasion in vitro and suppressed tumour growth in vivo. Downregulation of hsa_circ_0043278 led to the opposite results. Hsa_circ_0043278 expression was negatively correlated with that of miR-455-3p. In addition, mechanistic investigation proved that hsa_circ_0043278 directly bound to miR-455-3p and regulated EI24 and NF-κB expression in BC cells.Conclusion Hsa_circ_0043278 acts as a tumour suppressor gene in BC through the hsa_circ_0043278/miR-455-3p/EI24 axis and may be regarded as a new prognostic predictor or potential therapeutic target in BC.
研究领域:
肿瘤学
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分子生物学
·
遗传学
✓ 已验证
DOI
PubMed
高影响因子
Nature Communications
·
2025
·Vol. 16
·期 1
·1-19
·
IF 15.7
Zhang Min,Zhang Bin,Liu Chengli,Preußner Marco,Ayachit Megha,Li Weiming,Huang Yafei,Liu Deyi,He Quanwei,Emmerichs Ann-Kathrin,Meinke Stefan,Chen Shu,Wang Lin,Zheng Liduan,Li Qiubai,Huang Qin,Haltenhof Tom,Gao Ruoxi,Qin Xianan,Cheng Aifang,Wei Tianzi,Yu Li,Schubert Mario,Gao Xin,Li Mingchang,Heyd Florian
Mammals tightly regulate their core body temperature, yet how cells sense and respond to small temperature changes remains incompletely understood. Here, we discover RNA G-quadruplexes (rG4s) as key thermosensors enriched near splice sites of cold-repressed exons. These thermosensing RNA structures, when stabilized, mask splice sites, reducing exon inclusion. Specifically, rG4s near splice sites of a cold-repressed poison exon in the neuroprotective RBM3 are stabilized at low temperatures, leading to exon exclusion. This enables evasion of nonsense-mediated decay, increasing RBM3 expression at cold. Importantly, stabilizing rG4 through increasing intracellular potassium with an FDA-approved potassium channel blocker, mimics the hypothermic effect on alternative splicing, thereby increasing RBM3 expression, leading to RBM3-dependent neuroprotection in a mouse model of subarachnoid hemorrhage. Our findings unveil a mechanism how mammalian RNAs directly sense temperature and potassium perturbations, integrating them into gene expression programs. This opens new avenues for treating diseases arising from splicing defects and disorders benefiting from therapeutic hypothermia, especially hemorrhagic stroke.
研究领域:
分子生物学
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神经科学
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RNA生物学
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温度生物学
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基因表达调控
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产品关联
DOI
PubMed
高影响因子
PLoS One
·
2023
·Vol. 18
·期 3
·e0268917
·
IF 3.7
Chen Zhang,Qingrong Qu,Yuming Yao,Xiaobo Fan,Guoqiu Wu
The hybridization chain reaction (HCR) is widely used for biosensing. However, HCR does not provide the required sensitivity. In this study, we reported a method to improve the sensitivity of HCR by dampening the cascade amplification. First, we designed a biosensor based on HCR, and an initiator DNA was used to trigger the cascade amplification. Optimization of the reaction was then performed, and the results showed that the limit of detection (LOD) for the initiator DNA was about 2.5 nM. Second, we designed a series of inhibitory DNAs to dampen the HCR cascade amplification, and DNA dampeners (50 nM) were applied in the presence of the DNA initiator (50 nM). One of the DNA dampeners (D5) showed the best inhibitory efficiency of greater than 80%. This was further applied at concentrations ranging from 0 nM to 10 nM to prohibit the HCR amplification caused by a 2.5 nM initiator DNA (the limit of detection for this initiator DNA). The results showed that 0.156 nM of D5 could significantly inhibit the signal amplification (p<0.05). Additionally, the limit of detection for the dampener D5 was 16 times lower than that for the initiator DNA. Based on this detection method, we achieved a detection limit as low as 0.625 nM for HCV-RNAs. In summary, we developed a novel method with improved sensitivity to detect the target designed to prohibit the HCR cascade. Overall, this method could be used to qualitatively detect the presence of single-stranded DNA/RNA.
研究领域:
分子生物学
·
医学检验
·
生物化学
✓ 已验证
DOI
PubMed
高影响因子
LANGMUIR
·
2022
·
IF 4.331
Feixia Peng,Xuelian Wang,Wenting Tao,Yao Chen,Yani Ma,Xueqin Ding
Four kinds of hydrophobic magnetic deep eutectic solvents (HMDESs) were prepared and applied to RNA extraction. Based on the HMDESs, a mechanical shaking-assisted liquid–liquid extraction (MSLLE) was developed for the extraction of RNA. Factors that influence the extraction, including the extraction time, temperature, volume of HMDES, buffer types, and pH, were evaluated. After the optimization of all conditions, the RNA extraction efficiency was 82.31 ± 0.02%. RNA can be extracted from complex samples and medicinal yeast by the method proposed in this work and can be recovered from the HMDESs after being extracted.
研究领域:
生物化学
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
Current Protocols
·
2023
·Vol. 3
·期 11
·e923
·
IF 0
Xuan Chen,Long Xie,Chenglong Zhang,Shen Tian,Zeyu Tang,Weiwei Tian,Patrick Lu,Xianbin Yang
Although small interfering RNA (siRNA) is a key player among gene inhibition therapeutics, there are many obstacles to the development of siRNA drugs due to inherent properties of oligonucleotides, including the unsatisfactory stability of unmodified siRNA, poor pharmacokinetic distribution, and the toxicity induced by off-target effects. To maximize treatment potency, chemical modification of siRNA has undoubtedly been the most successful strategy by far. Widely applied modifications include phosphorothioate linkages, 2′- O -methyl modifications, and 2′-fluoro modifications, among others. To extend the family of chemical modifications for oligonucleotides, 2′- O -cyanoethylated RNA analogs were developed through the replacement of the 2′-hydroxyl group with a 2′- O -cyanoethyl group (-OCH 2 CH 2 CN). This modification can provide several advantages over unmodified RNA, such as increased stability, improved binding affinity to complementary DNA or RNA strands, and resistance to degradation by cellular nucleases. The 2′- O -cyanoethyl-modified RNAs not only are applied in RNA silencing machinery but also act as research tools for studying RNA structure and function or for developing RNA-based diagnostics. Therefore, the efficient synthesis, deprotection, purification, and characterization of 2′- O -cyanoethylated RNAs deserves more attention. This protocol describes the chemical synthesis of 2′- O -cyanoethylated nucleotides and the solid-phase synthesis, deprotection, and purification of 2′- O -cyanoethylated RNAs. © 2023 Wiley Periodicals LLC. Basic Protocol 1 : Preparation of 6- N -dimethylformamidyl-5′- O -dimethoxytrityl-2′- O -cyanoethyl adenosine 3′-(2-cyanoethyl N , N -diisopropyl)phosphoramidite Basic Protocol 2 : Preparation of 4- N -acetyl-5′- O -dimethoxytrityl-2′- O -cyanoethyl cytidine 3′-(2-cyanoethyl N , N -diisopropyl)phosphoramidite Basic Protocol 3 : Preparation of 2- N -dimethylformamidyl-5′- O -dimethoxytrityl-2′- O -cyanoethyl guanine 3′-(2-cyanoethyl N , N -diisopropyl)phosphoramidite Basic Protocol 4 : Preparation of 5′- O -dimethoxytrityl-2′- O -2-cyanoethyl uridine 3′-(2-cyanoethyl N , N -diisopropyl)phosphoramidite Basic Protocol 5 : Solid-phase synthesis of 2′- O -cyanoethylated RNA analogs Basic Protocol 6 : Deprotection and purification of synthesized 2′- O -cyanoethyl-RNAs
研究领域:
分子生物学
·
生物化学
·
药物化学
·
RNA治疗学
·
有机合成
✓ 已验证
DOI
Cell Reports
·
2025
·
IF 6.9
Yixuan Feng,Yiming Wang,Tai Wang,Lingtong Liu
Plant-unique RNA polymerase IV (RNA Pol IV) governs the establishment of small RNA (sRNA)-directed DNA methylation (RdDM). In dicotyledon, elevated RdDM activity is often associated with embryogenesis; however, the loss of RdDM frequently results in indiscernible phenotypes. Here, we report that the absence of SlNRPD1 , encoding the largest subunit of RNA Pol IV, leads to diminished RdDM and abnormal embryogenesis in tomato ( Solanum lycopersicum ). Hypermethylation at pericentric transposable elements (TEs) and a burst of 21/22-nt siRNA from the distal and pericentric genes are observed in slnrpd1 embryos. The specific activation of endoribonuclease Dicer-like 2 ( SlDCL2b / c / d ) is correlated with 21/22-nt sRNA accumulation. Auxin and WUSCHEL-related homeobox (WOX) signaling gene expression is altered by mCHH hypomethylation, which may lead to defective embryos. Due to improper maturation, the slnrpd1 embryos cannot withstand post-harvest desiccation. This study provides insights into how DNA methylation regulates plant embryogenesis.
研究领域:
植物生物学
·
表观遗传学
·
分子遗传学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
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