Fluorescence sensor for simultaneously detecting hOGG1 and hAAG and detection method and application of fluorescence sensor

Abstract

The invention belongs to the technical field of biological analysis, and particularly relates to a fluorescence sensor for simultaneously detecting hOGG1 and hAAG and a detection method and application of the fluorescence sensor. The fluorescence sensor comprises a difunctional dumbbell probe, a signal probe 1 and a signal probe 2, the difunctional dumbbell probe comprises a T7 promoter region, and is modified with a 8-oxoG and a deoxyinosine; the 5'ends of the signal probes 1 and 2 are respectively modified with different fluorophores, and the 3'ends of thesignal probes 1 and 2 are respectively modified with different quenchers . It is proved for the first time that controllable T7 transcriptional activation cyclic cascade amplification detects human 8-oxo guanine DNA glycosylase (hOGG1) and human alkyl adenine DNA glycosylase (hAAG) at the same time at the single molecular level, the method has good specificity and sensitivity, the detection limit of hOGG1 is 3.52 * 10<-8> U / mu L, and the detection limit of hAG is 3.55 * 10<-7> U / mu L; and even the glycosylase activity can be quantitatively repaired at a single cell level.

Description

technical field [0001] The invention belongs to the technical field of biological analysis, in particular to a fluorescence sensor for simultaneously detecting hOGG1 and hAAG, a detection method and an application thereof. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily taken as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] The specific pairing of heterocyclic bases (i.e., A with T, G with C) in the DNA double helix is ​​crucial for the preservation and transmission of genetic information encoded in the human genome. However, the chemical structure of heterocyclic bases has a large number of nucleophilic and redox active sites, which are often subjected to various exogenous (for example, ultraviolet light, genotoxic chemicals...

AI Technical Summary

Problems solved by technology

However, gel-based radiometric assays suffer from the following disadvantages: hazardous radiation and time-consuming manipulation; ELISA requires expensive antibodies and can be underestimated due to sample loss during multi-step washes

However, the inventors found that achieving probe binding and signal amplification at the interface of biosensors inevitably leads to reduced binding efficiency and enzyme kinetics due to steric hindrance, variable chemical microenvironment, and crowded surface effects.

Although improvements in interface engineering of nanostructures can allow targets to maximize recognition efficiency, variability in surface micro / nanofabrication may significantly affect the quantification and reproducibility of targets in complex matrices

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