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The advent and progression of AR (augmented reality) technology, coupled with the emergence of AR hearing aid glasses, offer a novel opportunity for people with hearing impairments (PHI). This study aims to explore the intention of this population to employ AR hearing aid glasses as their choice of hearing aid device and the specific factors influencing their preference. This study utilized the partial least squares SEM (PLS-SEM) analytical method to create structural equation model for intentions of PHI to use AR glasses as hearing aids. Data were gathered from on-site experiences across multiple locations; a total of 189 valid questionnaires from individuals with varying degrees of hearing disabilities were used for statistical analysis. According to the data analysis results, we discovered that functionality quality, perceived interaction speed, and perceived usability significantly influence communication effectiveness. Further, communication effectiveness positively influences confidence and societal perception, and the latter has a positive impact on information. Both of these factors positively influence behavioral intention. Based on these findings, this study offers design recommendations for AR hearing aid glasses to cater to the specific needs of PHI, aiming to enhance their quality of life. Furthermore, this study provides pivotal insights for the prospective growth of this emerging industry.
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Magnaporthe oryzae is a rice blast pathogen that seriously threatens rice yield. Benzovindiflupyr is a succinate dehydrogenase inhibitor (SDHI) fungicide that effectively controls many crop diseases. Benzovindiflupyr has a strong inhibitory effect on M. oryzae; however, control of rice blast by benzovindiflupyr and risk of resistance to benzovindiflupyr are not well studied in this pathogen. In this study, six benzovindiflupyr-resistant strains were obtained by domestication induced in the laboratory. The MoSdhBH245D mutation was the cause of M. oryzae resistance to benzovindiflupyr, which was verified through succinate dehydrogenase (SDH) activity assays, molecular docking, and site-specific mutations. Survival fitness analysis showed no significant difference between the benzovindiflupyr-resistant and parent strains. Positive cross-resistance to benzovindiflupyr and other SDHIs and negative cross-resistance to azoxystrobin were observed. Therefore, the risk of benzovindiflupyr resistance in M. oryzae might be medium to high. It should be combined with other classes of fungicides (tebuconazole and azoxystrobin) to slow the development of resistance.
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Farmacorresistência Fúngica , Fungicidas Industriais , Mutação , Succinato Desidrogenase , Succinato Desidrogenase/genética , Succinato Desidrogenase/antagonistas & inibidores , Fungicidas Industriais/farmacologia , Farmacorresistência Fúngica/genética , Doenças das Plantas/microbiologia , Magnaporthe/efeitos dos fármacos , Magnaporthe/genética , Simulação de Acoplamento Molecular , Oryza/microbiologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Estrobilurinas/farmacologia , AscomicetosRESUMO
A CRISPR/Cas12a-coupled multiplexed strand displacement amplification (CMSDA) for the detection of miR155 has been developed. Non-specific amplification was avoided by designing a single-stranded DNA template with a hairpin structure. The detection target miR155 was used as a primer to initiate a multiple-strand displacement reaction to produce abundant ssDNA. ssDNA was recognized by the Cas12a/CrRNA binary complex, activating the trans-cleaving activity of Cas12a. The multiple-strand displacement reaction is more efficiently detected compared with a single-strand displacement reaction. The detection range is from 250 pM to 1 nM, and the limit of the detection is 6.5 pM. The proposed method showed a good applicability in complex serum environments, indicating that the method has a broad prospect for disease detection and clinical application. In addition, we designed a dual-cavity PCR tube, which realized one-tube detection of miRNA155 and avoided open-cap contamination.
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Sistemas CRISPR-Cas , MicroRNAs , MicroRNAs/análise , MicroRNAs/sangue , MicroRNAs/genética , Humanos , Sistemas CRISPR-Cas/genética , DNA de Cadeia Simples/química , DNA de Cadeia Simples/genética , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase/métodos , Proteínas de Bactérias , Endodesoxirribonucleases , Proteínas Associadas a CRISPRRESUMO
Fast, sensitive, and portable detection of genetic modification contributes to agricultural security and food safety. Here, we developed RPA-CRISPR/Cas12a-G-quadruplex colorimetric assays that can combine with intelligent recognition by deep learning algorithms to achieve sensitive, rapid, and portable detection of the CaMV35S promoter. When the crRNA-Cas12a complex recognizes the RPA amplification product, Cas12 cleaves the G-quadruplex, causing the G4-Hemin complex to lose its peroxide mimetic enzyme function and be unable to catalyze the conversion of ABTS2- to ABTS, allowing CaMV35S concentration to be determined based on ABTS absorbance. By utilizing the RPA-CRISPR/Cas12a-G4 assay, we achieved a CaMV35S limit of detection down to 10 aM and a 0.01 % genetic modification sample in 45 min. Deep learning algorithms are designed for highly accurate classification of color results. Yolov5 objective finding and Resnet classification algorithms have been trained to identify trace (0.01 %) CaMV35S more accurately than naked eye colorimetry. We also coupled deep learning algorithms with a smartphone app to achieve portable and rapid photo identification. Overall, our findings enable low cost ($0.43), high accuracy, and intelligent detection of the CaMV35S promoter.
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Sistemas CRISPR-Cas , Colorimetria , Aprendizado Profundo , Quadruplex G , Colorimetria/métodos , Sistemas CRISPR-Cas/genética , Regiões Promotoras Genéticas , Proteínas Associadas a CRISPR/genética , Proteínas Associadas a CRISPR/metabolismo , Limite de Detecção , Proteínas de Bactérias/genética , EndodesoxirribonucleasesRESUMO
Intracellular glucose detection is crucial due to its pivotal role in metabolism and various physiological processes. Precise glucose monitoring holds significance in diabetes management, metabolic studies, and biotechnological applications. In this study, we developed an innovative and expedient cell-permeable nanoreactor for intracellular glucose based on surface-enhanced Raman scattering (SERS). The nanoreactor was designed with gold nanoparticles (AuNPs), which were engineered with glucose oxide (GOx) and a H2O2-responsive Raman reporter 2-mercaptohydroquinone (2-MHQ). The interaction between 2-MHQ and H2O2 generated by glucose and GOx could simultaneously induce the appearance in the peak at 985 cm-1. Our results showed excellent performance in detecting glucose within the concentration range from 0.1 µM to 10 mM, with a low detection limitation of 14.72 nM. In addition, the glucose distribution in single HeLa cells was evaluated by real time SERS mapping. By combining noble metal particles and natural oxidases, the nanoreactor possesses both Raman activity and enzymatic functionality, thus enables sensitive glucose detection and facilitates imaging at a single cell level, which offers an insightful monitoring of cellular processes.
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Glucose , Ouro , Nanopartículas Metálicas , Análise Espectral Raman , Análise Espectral Raman/métodos , Humanos , Células HeLa , Ouro/química , Nanopartículas Metálicas/química , Glucose/análise , Glucose/metabolismo , Peróxido de Hidrogênio/análise , Peróxido de Hidrogênio/química , Glucose Oxidase/química , Glucose Oxidase/metabolismoRESUMO
Hypoxia is a hallmark of cancer development. However, the molecular mechanisms by which hypoxia promotes tumor metastasis are not fully understood. In this study, we demonstrate that hypoxia promotes breast cancer metastasis through suppression of ΔNp63α in a HIF1α-independent manner. We show that hypoxia-activated XBP1s forms a stable repressor protein complex with HDAC2 and EZH2 to suppress ΔNp63α transcription. Notably, H3K27ac is predominantly occupied on the ΔNp63 promoter under normoxia, while H3K27me3 on the promoter under hypoxia. We show that XBP1s binds to the ΔNp63 promoter to recruit HDAC2 and EZH2 in facilitating the switch of H3K27ac to H3K27me3. Pharmacological inhibition or the knockdown of either HDAC2 or EZH2 leads to increased H3K27ac, accompanied by the reduced H3K27me3 and restoration of ΔNp63α expression suppressed by hypoxia, resulting in inhibition of cell migration. Furthermore, the pharmacological inhibition of IRE1α, but not HIF1α, upregulates ΔNp63α expression in vitro and inhibits tumor metastasis in vivo. Clinical analyses reveal that reduced p63 expression is correlated with the elevated expression of XBP1, HDAC2, or EZH2, and is associated with poor overall survival in human breast cancer patients. Together, these results indicate that hypoxia-activated XBP1s modulates the epigenetic program in suppression of ΔNp63α to promote breast cancer metastasis independent of HIF1α and provides a molecular basis for targeting the XBP1s/HDAC2/EZH2-ΔNp63α axis as a putative strategy in the treatment of breast cancer metastasis.
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Neoplasias da Mama , Proteína Potenciadora do Homólogo 2 de Zeste , Epigênese Genética , Histona Desacetilase 2 , Subunidade alfa do Fator 1 Induzível por Hipóxia , Proteínas Supressoras de Tumor , Proteína 1 de Ligação a X-Box , Humanos , Neoplasias da Mama/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteína 1 de Ligação a X-Box/metabolismo , Proteína 1 de Ligação a X-Box/genética , Histona Desacetilase 2/metabolismo , Histona Desacetilase 2/genética , Feminino , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas Supressoras de Tumor/genética , Animais , Linhagem Celular Tumoral , Metástase Neoplásica , Camundongos , Regulação Neoplásica da Expressão Gênica , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Hipóxia Celular/genéticaRESUMO
Chaperonins, which contain t-complex polypeptide 1 (CCT), are critical for correct protein folding to generate stable and functional protein conformations, which are important for cell growth and survival. However, little is known about the expression and prognostic significance of CCT8 (subunit 8 of the CCT complex chaperonin) in lung cancer. In this study, we demonstrated that CCT8 expression is frequently increased in human lung cancer. Survival analysis indicated that CCT8 expression is closely correlated with inferior overall survival in lung adenocarcinoma (LUAD), but not in lung squamous carcinoma (LUSC). Subsequently, ectopic expression of CCT8 facilitated cell migration and tumor metastasis, and vice versa. Mechanistically, CCT8 interacted and activated ATK. Inhibition of AKT suppressed CCT8-induced cell migration and tumor metastasis. Our findings support CCT8 as a biomarker for LUAD prognosis and as a target for LUAD therapy.
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Palladium is a vital commodity in the industry. To guarantee a stable supply in the future, it is imperative to adopt more effective recycling practices. In this proof-of-concept study, we explore the potential of electrodialysis to enhance the palladium concentration in a residual solution of palladium recycling, thus promoting higher recovery rates. Experiments were conducted using an industrial hydrochloric acid solution containing around 1000 mg/L of palladium, with a pH below 1. Two sets of membranes, Selemion AMVN/CMVN and Fujifilm Type 12 AEM/CEM, were tested at two current levels. The Fujifilm membranes, which are designed for low permeability of water, show promising results, recovering around 40% of palladium within a two-hour timeframe. The Selemion membranes were inefficient due to excessive water transport. All membranes accumulated palladium in their structures. Anion-exchange membranes showed higher palladium accumulation at lower currents, while cation-exchange membranes exhibited increased palladium accumulation at higher currents. Owing to the low concentration of palladium and the presence of abundant competing ions, the current efficiency remained below 2%. Our findings indicate a strong potential for augmenting the palladium stage in industrial draw solutions through electrodialysis, emphasizing the importance of membrane properties and process parameters to ensure a viable process. Beyond the prominent criteria of high permselectivity and low resistance, minimizing the permeability of water within IEMs remains a key challenge to mitigating the efficiency loss associated with uncontrolled mixing of the electrolyte solution.
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Herein, we integrated CRISPR/Cas12a with primer-assisted rolling circle amplification (PARCA) to specifically detect EGFR 19 from the genome. We fused the method into fluorescent and electrochemical detection systems forming a stable and sensitive dual-signal sensing platform. The fluorescent detection system stably detected EGFR 19 in a linear range from 500 fM to 10 nM with an ultra-low background signal. The electrochemical detection system possessed a detection limit as low as 42 aM due to the introduction of nanomaterial UIO-66-NH2. The dual-signal sensing platform showed superior performance in complex serum samples and real cell genomes and provided a flexible and dynamic approach for the ultra-sensitive detection of EGFR 19.
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Técnicas Biossensoriais , Sistemas CRISPR-Cas , Sistemas CRISPR-Cas/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Corantes , Receptores ErbB/genética , Técnicas Biossensoriais/métodosRESUMO
[Purpose/Significance] Under the influence of various factors such as the pandemic, oil prices, and economic conditions, the global automotive industry has entered a period of downturn. Therefore, how to activate market potential and increase consumers' willingness to purchase cars has become an important research topic. Unlike many other products, test drives play a significant role in the car-buying process. [Method/Procedure] This study employs a questionnaire survey to explore how consumer perceptions of product aesthetics, space quality, and service quality during their initial visit to an automobile 4S store influence their test drive intention through two dimensions of brand image: symbolic and experiential. A structural equation model is used to establish a test drive intention impact model incorporating these dimensions. [Results/Conclusions] The study found that brand image, both symbolic and experiential, plays a significant mediating role in enhancing potential consumers' test drive intentions. Space image had the most significant impact on brand image. Although product aesthetics did not directly affect brand image experientially, they remained an important factor in enhancing brand image symbolically. [Contribution/Value] The results of this study can provide insights for automotive brand managers, automobile 4S store designers, and others aiming to promote the sustainable development of automotive consumption.
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Accurate detection of cancer-associated mRNAs is beneficial to early diagnosis and potential treatment of cancer. Herein, for the first time, we developed a novel CRISPR/Cas12a-powered electrochemical/fluorescent (EC/FL) dual-mode controlled-release homogeneous biosensor for mRNA detection. A functionalized ssDNA P2-capped Fe3O4-NH2 loaded with methylene blue (P2@MB-Fe3O4-NH2) was synthesized as the signal probe, while survivin mRNA was chosen as the target RNA. In the presence of the target mRNA, the nicking endonuclease-mediated rolling circle amplification (NEM-RCA) was triggered to produce significant amounts of ssDNA, activating the collateral activity of Cas12a toward the surrounding single-stranded DNA. Thus, the ssDNA P1 completely complementary to ssDNA P2 was cleaved, resulting in that the ssDNA P2 bio-gate on Fe3O4-NH2 could not be opened due to electrostatic interactions. As a result, there was no or only a little MB in the supernatant after magnetic separation, and the measured EC/FL signal was exceedingly weak. On the contrary, the ssDNA P2 bio-gate was opened, enabling MB to be released into the supernatant, and generating an obvious EC/FL signal. Benefiting from the accuracy of EC/FL dual-mode cross-verification, high amplification efficiency, high specificity of NEM-RCA and CRISPR/Cas12a, and high loading of mesoporous Fe3O4-NH2 on signal molecules, the strategy shows aM-level sensitivity and single-base mismatch specificity. More importantly, the practical applicability of this dual-mode strategy was confirmed by mRNA quantification in complex serum environments and tumor cell lysates, providing a new way for developing a powerful disease diagnosis tool.
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Técnicas Biossensoriais , Sistemas CRISPR-Cas , Sistemas CRISPR-Cas/genética , Preparações de Ação Retardada , RNA Mensageiro/genética , RNA , Corantes , DNA de Cadeia Simples/genética , Endonucleases , Inibidores de Serina ProteinaseRESUMO
The selective separation of metal species from various sources is highly desirable in applications such as hydrometallurgy, water treatment, and energy production but also challenging. Monovalent cation exchange membranes (CEMs) show a great potential to selectively separate one metal ion over others of the same or different valences from various effluents in electrodialysis. Selectivity among metal cations is influenced by both the inherent properties of membranes and the design and operating conditions of the electrodialysis process. The research progress and recent advances in membrane development and the implication of the electrodialysis systems on counter-ion selectivity are extensively reviewed in this work, focusing on both structure-property relationships of CEM materials and influences of process conditions and mass transport characteristics of target ions. Key membrane properties, such as charge density, water uptake, and polymer morphology, and strategies for enhancing ion selectivity are discussed. The implications of the boundary layer at the membrane surface are elucidated, where differences in the mass transport of ions at interfaces can be exploited to manipulate the transport ratio of competing counter-ions. Based on the progress, possible future R&D directions are also proposed.
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MicroRNAs (miRNAs) play a crucial role in the regulation of gene expression and have been implicated in many diseases. Herein, we develop a target triggered exponential rolling-circle amplification coupling with CRISPR/Cas12a (T-ERCA/Cas12a) system, which can achieve the ultrasensitive detection with simple operation and no annealing procedure. In this assay, T-ERCA combines the exponential amplification with rolling-circle amplification by introducing a dumb-bell probe with two enzyme recognition sites. miRNA-155 targets are activators that trigger exponential rolling circle amplification to produce large amounts of ssDNA, which is then recognized by CRISPR/Cas12a for further amplification. Compared with single EXPAR or RCA combined with CRISPR/Cas12a, this assay shows higher amplification efficiency. Therefore, benefiting from the excellent amplification effect of T-ERCA and the high recognition specificity of CRISPR/Cas12a, the proposed strategy shows a wide detection range from 1 fM to 5 nM with a LOD (limit of detection) down to 0.31 fM. Moreover, it shows good application ability for assessing miRNA levels in different cells, indicating that the T-ERCA/Cas12a may provide a new guidance for molecular diagnosis and clinical practical application.
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Técnicas Biossensoriais , MicroRNAs , MicroRNAs/genética , MicroRNAs/metabolismo , Sistemas CRISPR-Cas , Técnicas de Amplificação de Ácido Nucleico/métodos , DNA de Cadeia Simples , Bioensaio/métodos , Técnicas Biossensoriais/métodosRESUMO
An improved electrochemical sensor has been developed for sensitive detection of the p53 gene based on exponential amplification reaction (EXPAR) and CRISPR/Cas12a. Restriction endonuclease BstNI is introduced to specifically identify and cleave the p53 gene, generating primers to trigger the EXPAR cascade amplification. A large number of amplified products are then obtained to enable the lateral cleavage activity of CRISPR/Cas12a. For electrochemical detection, the amplified product activates Cas12a to digest the designed block probe, which allows the signal probe to be captured by the reduced graphene oxide-modified electrode (GCE/RGO), resulting in an enhanced electrochemical signal. Notably, the signal probe is labeled with large amounts of methylene blue (MB). Compared with traditional endpoint decoration, the special signal probe effectively amplifies the electrochemical signals by a factor of about 15. Experimental results show that the electrochemical sensor exhibits wide ranges from 500 aM to 10 pM and 10 pM to 1 nM, as well as a relatively low limit detection of 0.39 fM, which is about an order of magnitude lower than that of fluorescence detection. Moreover, the proposed sensor shows reliable application capability in real human serum, indicating that this work has great prospects for the construction of a CRISPR-based ultra-sensitive detection platform.
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Sistemas CRISPR-Cas , Genes p53 , Humanos , Primers do DNA , Eletrodos , FluorescênciaRESUMO
Despite numerous publications on membrane materials and the fabrication of thin-film composite (TFC) membranes for CO2 separation in recent decades, the effects of porous supports on TFC membrane performance have rarely been reported, especially when humid conditions are concerned. In this work, six commonly used porous supports were investigated to study their effects on membrane morphology and the gas transport properties of TFC membranes. Two common membrane materials, Pebax and poly(vinyl alcohol) (PVA), were employed as selective layers to make sample membranes. The fabricated TFC membranes were tested under humid conditions, and the effect of water vapor on gas permeation in the supports was studied. The experiments showed that all membranes exhibited notably different performances under dry or humid conditions. For polyacrylonitrile (PAN) and poly(ether sulfones) (PESF) membranes, the water vapor easily condenses in the pores of these supports, thus sharply increasing the mass transfer resistance. The effect of water vapor is less in the case of polyvinylidene difluoride (PVDF) and polysulfone (PSF), showing better long-term stability. Porous supports significantly contribute to the overall mass transfer resistance. The presence of water vapor worsens the mass transfer in the porous support due to the pore condensation and support material swelling. The membrane fabrication condition must be optimized to avoid pore condensation and maintain good separation performance.
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DNA methylation is considered as a potential cancer biomarker. The evaluation of DNA methylation level will contribute to the prognosis and diagnosis of cancer. Herein, we propose a novel assay based on endonuclease-assisted protospacer adjacent motif (PAM)-free recombinase polymerase amplification coupling with CRISPR/Cas12a (E-PfRPA/Cas) for sensitive detection of DNA methylation. The methylation-sensitive restriction enzyme (MSRE) is first used to selectively digest unmethylated DNA, while the methylated target remains structurally intact. Therefore, the methylated target can initiate the RPA reaction to generate a large amount of double-stranded DNA (dsDNA). To avoid the dependence of PAM site of CRISPR/Cas12a, one of the RPA primers is designed with 5'-phosphate terminuses. After treating with Lambda, the sequence with 5'-phosphate modification will be degraded, leaving the single-stranded DNA (ssDNA). The CRISPR/Cas12a can accurately locate ssDNA without PAM, then initiating its trans-cleavage activity for further signal amplification. Meanwhile, non-specific amplification can be also avoided under Lambda, effectively filtering the detection background. Benefiting from the specificity of MSRE, the high amplification efficiency of Lambda-assisted RPA, and the self-amplification effect of CRISPR/Cas, the E-PfRPA/Cas assay shows outstanding sensitivity and selectivity, and as low as 0.05% of methylated DNA can be distinguished. Moreover, the lateral flow assay is also introduced to exploit the point-of-care diagnostic platform. Most importantly, the proposed method shows high sensitivity for determination of genomic DNA methylation from cancer cells, indicating its great potential for tumor-specific gene analysis.
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Proteínas Associadas a CRISPR , Proteínas Associadas a CRISPR/genética , Proteínas Associadas a CRISPR/metabolismo , Endonucleases/genética , Endonucleases/metabolismo , Sistemas CRISPR-Cas/genética , Recombinases/genética , Recombinases/metabolismo , Endodesoxirribonucleases/genética , Endodesoxirribonucleases/metabolismo , Metilação de DNA , Proteínas de Bactérias/genética , DNA de Cadeia Simples/genética , DNA/análise , FosfatosRESUMO
Background: Accumulating evidences have revealed that the abnormal N6-methyladenosine (m6A) modification is closely associated with the occurrence, development, progression and prognosis of cancer. It is noteworthy that m6A modification is widely existed in circRNAs and found its key biological functions in regulating circRNAs metabolism. However, the role of m6A modified circRNAs in colorectal cancer (CRC) remains unknown. To better understand the role of circRNAs in the pathogenesis of CRC, we focus on the relationship between m6A-modified circRNAs and their parental genes. Methods: Arraystar m6A-circRNA epitranscriptomic microarray was used to identify differentially m6A modified circRNAs between CRC and the control group. In addition, TCGA-COAD and GSE106582 cohort were used to identify differentially expressed mRNAs. In this study, we screened the parental genes for which both circRNAs and mRNAs were down-regulated further to analyze, including gene expression, survival prognosis, enrichment analysis. Additionally, Western Blotting was used to further validate the role of the parental gene in CRC. Results: We found that 1405 significantly downregulated circRNAs in CRC by our microarray data. Moreover, we obtained 113 parental genes for which both circRNAs and mRNAs were down-regulated to analyze the relationship with the prognosis of CRC based on TCGA-COAD cohort. And we identified nine potential prognostic genes, including ABCD3, ABHD6, GAB1, MIER1, MYOCD, PDE8A, RPS6KA5, TPM1 and WDR78. And low expression of these genes was associated with poor survival prognosis of the patients with CRC. In addition, we found that TPM1 is downregulated in CRC by western blotting experiment. And the calcium-signaling pathway may involve the process of the CRC progression. Conclusions: We identified nine potential prognostic genes, after analyzed the relationship between the parental genes of m6A modified circRNAs and the progression of CRC. Above all, our study further validated TPM1 can serve as a potentail signature for CRC patients.
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A novel fluorescent biosensor was proposed for detecting the CaMV 35S promoter in genetically modified organisms (GMOs). It was based on a proximity extension mediated multiple cascade strand displacement amplification connected with CRISPR/Cpf 1 (termed PE-MC/SDA-CRISPR/Cpf1). In this protocol, the CaMV 35S was recognized by proximity reaction in the presence of two adjacent primer probes. The proximity extension further triggered the multiple cascade strand displacement amplification (MC/SDA), generating a mass of ssDNA. The products compelled the trans-cleavage activity of CRISPR/Cpf 1, so as to cleave nearby ssDNA-FQ reporters and generate a strong fluorescent signal. The ingenious three-link combination design allowed the CaMV 35S a low background interference. And the MC/SDA combined with CRISPR/Cpf 1 dramatically improved the detection sensitivity. Under optimized conditions, the detection linear range of ultrasensitive fluorescent biosensor for CaMV 35S was from 50 fM to10 pM and 10 pM-500 pM, along with the limit of detection (LOD) down to 14.4 fM. The sensing platform also had excellent performance in the assay of selectivity and real samples. Therefore, the method earned great application potential for transgenic crops.
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Técnicas Biossensoriais , Técnicas de Amplificação de Ácido Nucleico , Técnicas Biossensoriais/métodos , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico/métodos , Regiões Promotoras GenéticasRESUMO
Inborn errors of metabolism (IEMs) are strongly related to abnormal growth and development in newborns and can even result in death. In total, 94,648 newborns were enrolled for expanded newborn screening using tandem mass spectrometry (MS/MS) from 2016 to 2020 at the Neonatal Disease Screening Center of the Maternal and Child Health Hospital in Shaoyang City, China. A total of 23 confirmed cases were detected in our study with an incidence rate of 1:4,115. A total of 10 types of IEM were identified, and the most common IEMs were phenylalanine hydroxylase deficiency (PAHD; 1:15,775) and primary carnitine deficiency (PCD; 1:18,930). Mutations in phenylalanine hydroxylase (PAH) and SLC22A5 were the leading causes of IEMs. To evaluate the application effect of artificial intelligence (AI) in newborn screening, we used AI to retrospectively analyze the screening results and found that the false-positive rate could be decreased by more than 24.9% after using AI. Meanwhile, a missed case with neonatal intrahepatic cholestasis citrin deficiency (NICCD) was found, the infant had a normal citrulline level (31 µmol/L; cutoff value of 6-32 µmol/L), indicating that citrulline may not be the best biomarker of intrahepatic cholestasis citrin deficiency. Our results indicated that the use of AI in newborn screening could improve efficiency significantly. Hence, we propose a novel strategy that combines expanded neonatal IEM screening with AI to reduce the occurrence of false positives and false negatives.
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High temperatures are a major concern that limit rice germination and plant growth. Although previous studies found that melatonin can promote seed germination, the physiological regulation mechanism by which exogenous melatonin mediates high temperature tolerance during rice seed germination is still largely unknown. In order to overcome these challenges, the present study investigates the effects of melatonin on the characteristics of rice seed germination as well as on antioxidant properties, under different high temperature conditions. The results show that 100 µM melatonin seed-soaking treatment under high temperature conditions effectively improves the germination potential, the germination index, and the vigor index of rice seeds; increases the length of the shoot and the root; improves the activity of the antioxidant enzymes; and significantly reduces the malondialdehyde content. The gray relational grade of the shoot peroxidase activity and the melatonin soaking treatment was the highest, which was used to evaluate the effect of melatonin on the heat tolerance of rice. The subordinate function method was used to comprehensively evaluate the tolerance, and the results show that the critical concentration of melatonin is 100 µM, and the critical interactive treatment is the germination at 38 °C and followed by the recovery at 26 °C for 1 day + 100 µM. In conclusion, 100 µM of melatonin concentration improved the heat resistance of rice seeds by enhancing the activity of the antioxidant enzymes.