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Biosensor for simultaneously detecting multiple methyltransferases of DNA (deoxyribonucleic acid) as well as detection method and application of biosensor

A methyltransferase, biosensor technology, applied in biochemical equipment and methods, microbial determination/inspection, biomaterial analysis, etc. The effect of the cost of experiments

Inactive Publication Date: 2021-02-05
SHANDONG NORMAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But there are some limitations when applying them to the simultaneous detection of multiple DNA methyltransferases (MTases), these methods require additional substrate probes and fluorophores of different colors, which complicates the experimental procedure and increases the cost

Method used

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  • Biosensor for simultaneously detecting multiple methyltransferases of DNA (deoxyribonucleic acid) as well as detection method and application of biosensor
  • Biosensor for simultaneously detecting multiple methyltransferases of DNA (deoxyribonucleic acid) as well as detection method and application of biosensor
  • Biosensor for simultaneously detecting multiple methyltransferases of DNA (deoxyribonucleic acid) as well as detection method and application of biosensor

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Experimental program
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Embodiment

[0074] Preparation of dumbbell probes: 10 μl of annealing reaction system including 2 μl of 100 μmol per liter of dumbbell-shaped probe, 1.5 μl of 10×T4 DNA ligase buffer (500 mmol per liter of trihydroxy Methylaminomethane-hydrochloric acid (Tris-HCl), 100 millimoles per liter of magnesium chloride (MgCl 2 ), 100 mmol per liter of dithiothreitol (DTT) and 6.5 microliters of water. React at 95°C for 5 minutes, then slowly cool to room temperature. 100 units of T4 DNA ligase and 10 microliters of the above mixture were used to form a 15 microliter reaction system, and the DNA ligase reaction was carried out overnight at 16°C, and then inactivated at 65°C for 10 minutes. Subsequently, in 1×NEBuffer 1 containing 10 units of Exo I (exonuclease I), 10 units of Exo III (exonuclease III), 1×NEBuffer 1 (1,3-bis((trihydroxy Methyl)methylamino)propane-hydrochloric acid buffer, 10 mmol per liter of magnesium chloride (MgCl 2 ), 1 mmol per liter of dithiothreitol (DTT), pH 7.0), 1 × Ex...

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Abstract

The invention provides a biosensor for simultaneously detecting multiple methyltransferases of DNA as well as a detection method and an application of the biosensor, and belongs to the technical fieldof molecular detection. The biosensor at least comprises a dumbbell type probe, an annular template and a signal probe. The dumbbell type probe with the double-signal amplification function has two methylase recognition sites, so that the double-signal amplification technology based on the dumbbell type probe is matched with the annular template and the signal probe simultaneously to achieve ultra-sensitive detection of methylase, and therefore, the biosensor has huge potential in disease diagnosis and drug discovery, and has good practical application value in biomedical research.

Description

technical field [0001] The invention belongs to the technical field of molecular detection, and in particular relates to a biosensor for simultaneously detecting multiple DNA methyltransferases, 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 abnormal expression of DNA methyltransferases (MTases) is closely related to many diseases, and the abnormal activity of DNA methyltransferases (MTases) may destroy the normal DNA methylation pattern and induce a variety of cancers, including lung cancer, breast cancer, prostate cancer, colon cancer, liver cancer, kidney cancer, cervical cancer, thyroid cancer, retinoblastoma, and he...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/682C12Q1/6825C12Q1/48G01N21/64
CPCC12Q1/682C12Q1/6825C12Q1/48G01N21/6428G01N2333/91017
Inventor 张春阳李玥颖孙淑丽田小锐
Owner SHANDONG NORMAL UNIV
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