Articles
Publication Date: 2025
Analytical Methods (17599679)17(45)pp. 9244-9253
In recent years, thioflavin T (ThT) has been applied as an intercalating fluorescence probe in the development of biosensors. One approach is based on monitoring the fluorescent signal of ThT upon binding to a G-quadruplex (GQ). GQ is a structure that is formed from guanine-rich sequences. The sequences may be synthesized during isothermal or cyclic amplifications. The ThT-based fluorescent signal generation has been combined with various isothermal amplification methods. However, to the best of our knowledge, it has not yet been studied in combination with cyclic amplifications, i.e., polymerase chain reaction (PCR). PCR has benefits that has led to success in many commercial and FDA-approved diagnostic kits; however, the traditional qPCR methods that apply TaqMan technology suffer from challenges such as high price and dependence on sophisticated instruments. The current commercial qPCR kits are limited in their wider application by general public health authorities upon their price and availability. Thus, developing new amplification-based nucleic acid tests (NATs) was portrayed to be necessary to facilitate an improvement in public health. Here, we present a fluorimetric analysis of PCR products upon incorporating thioflavin T into the synthesized GQ as a new readout system for detecting a viral RNA. The PCR primers were designed with a reverse complementary sequence of a GQ. The system's efficiency was validated in one-pot, one-step reactions for detecting the presence of a model virus, i.e., SARS-CoV-2 genome in samples taken from individuals with various viral loads.
Publication Date: 2025
Scientific Reports (20452322)15(1)
Emerging evidence supports the role of type 2 diabetes (T2D) mellitus as a risk factor for cancer progression. In this study, we investigated and identified biomarkers related to diabetes and pancreatic ductal adenocarcinoma (PDAC) using systems biology to understand better the molecular landscape of PDAC and its connections with T2D. RNA-seq data related to blood samples of diabetes and pancreatic cancer were analyzed using bioinformatics tools in the Galaxy platform. After differential expression analysis using the DESeq2, the co-expression network associated with T2D and PDAC data was reconstructed using the WGCNA. Then, by visualizing the protein-protein interaction network in modules specifically related to T2D and PDAC, the key genes involved in these two diseases were identified, and their interaction network with long non-coding RNAs was reconstructed. Finally, the results of bioinformatics analysis were verified by qPCR in four groups, including T2D, PDAC, PDAC-T2D, and control groups. In this study, 1905 and 18,558 genes with significant differential expression were identified in the data of T2D and PDAC, respectively (|logFC| > 0.58, adj. p value < 0.05). The WGCNA showed 32 and 20 co-expression modules in diabetes and pancreatic cancer data, respectively. Among these, 303 genes were co-expressed, related to diabetes and pancreatic cancer. Based on the protein-protein interaction pattern, five hub genes were identified (using the CytoHubba Cytoscape plugin and the Maximal Clique Centrality (MCC) parameter). Finally, the co-expression network was reconstructed between these five genes and other lncRNAs. The qPCR showed that the expression of the CEBPZ gene was significantly increased in the blood samples of the diabetic (log2FC = 1.163, adj. p value = 0.0006), pancreatic cancer (log2FC = 3.22, adj. p value < 0.0001), and pancreatic cancer-diabetic (log2FC = 2.73, adj. p value < 0.0001) groups compared to the control group. For the first time, this study suggested that CEBPZ expression may serve as a diagnostic biomarker for assessing PDAC in individuals with T2D, given its differential expression in this specific cohort. © The Author(s) 2025.
