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Cell signal transduction activity detection method based on tetrahedral DNA nano material

A nanomaterial and cell signaling technology, applied in the field of biomedical detection, to achieve the effect of simple synthesis method, good reproducibility and flexible design

Pending Publication Date: 2022-03-01
THE FIRST AFFILIATED HOSPITAL OF FUJIAN MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are few reports on the application of TDNs to specific processes of cell signal transduction such as transcription factor activation and the related cell phenotype signals regulated by them.

Method used

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  • Cell signal transduction activity detection method based on tetrahedral DNA nano material
  • Cell signal transduction activity detection method based on tetrahedral DNA nano material
  • Cell signal transduction activity detection method based on tetrahedral DNA nano material

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Such as figure 1As shown, the tetrahedron DNA nanomaterial TDNS and replacement chain T have been designed, the tetrahedron DNA nanomaterial TDNS is made of five single-stranded DNA, and the five single-stranded DNA are TDNS-A, TDNS-B, TDNS. -C, TDNS-D and TDNS-cDNA, where

[0037] The 5 'end fragment of TDNS-A has a continuous series repeating sequence of the above-mentioned specific DNA binding motif reverse complement.

[0038] The 5 'end of TDNS-A has a fluorescent group FAM.

[0039] TDNS-CDNA is completely complemented with the above-mentioned continuous series repeat sequences,

[0040] The replacement chain T is complementary pair with the above-described continuous series repeat sequence, and has an additional sequence at its 5 'end to make the stability of the double-stranded structure formed by the conversion chain T and TDNS-A greater than the double formed by TDNS-CDNA and TDNS-A. The stability of the chain structure is, and the 5 'end of the chain T is BHQ1 ha...

Embodiment 2

[0047] Example 2 TGF-β signal transduction activity detection

[0048] The TGF-β signaling pathway plays an important role in early embryonic development and tissue organ formation, immunological supervision, and organizational repair and adult steady-state balance. TGF-β signal transduction abnormality may result in a variety of diseases, such as embryonic development, tumor, tissue fibrosis, cardiovascular disease, and immunological diseases. In this example, the mouse embryonic fibroblast (MEF) is used as an object, and the activity of TGF-β signaling path transcription factor SMAD 2 / 3 is detected by a new method based on TDNS. When the TGF-β signal path is activated, the phosphorylated transcription factor SMAD 2 / 3 can be combined with SMAD4 to form a heterologous triple composite. Such a complex can be transferred to the nucleus and combined with a unique DNA motif in the genome to adjust the expression of the TGF-β signal downstream target gene. Therefore, the present embodi...

Embodiment 3

[0063] Example 3 Oxygen-deficient signal transduction activity detection

[0064]This embodiment selects the hypoxia signal transduction passage in the u-87 mg of glioma cells as a detection object. Hypoxia Inducible Factor-α, HiF-α) is a transcription factor that is widely present in mammals and humans under hypoxia conditions, and is a key factor that responds to hypoxia complica. The specific DNA binding motif of the hypoxic transcription factor HIF-α is NCGTGN, wherein n represents any one of four bases of A, T, C, and G. Thus, this embodiment provides TDNS2 having a 5 (optional range of 5-10) having NCGTGN and a TDNS2 containing the FAM-labeled repeating sequence. As described above, the detection system comprises TDNS2 and a chain T2, wherein TDNS2 is formed by constituting chains TDNS2-A, B, CDNA (TDNS2 self-assembly conditions as in Example 1) by constituting chains TDNS2-A, B, CDNA (TDNS2 self-assembly conditions in Example 1) TDNS1), the relevant sequence is shown in Tab...

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Abstract

The invention discloses a cell signal transduction activity detection method based on a tetrahedral DNA nano-material, which comprises a specific DNA binding sequence of a transcription factor capable of characterizing signal transduction activity to be detected and a fluorescent label, when a target transcription factor is bound with TDNs, a fluorophore and a quenching group are prevented from approaching, and when the target transcription factor is bound with TDNs, the signal transduction activity to be detected is detected. Therefore, the transcription factor activity can be analyzed by detecting the fluorophore. The method is simple, convenient and efficient, qualitative, quantitative and imaging analysis can be synchronously completed, and the operation time is short. In addition, the synthesis method of the tetrahedral DNA nano material is simple and convenient, has good reproducibility, can be flexibly designed according to target molecules to be detected, and has strong universality.

Description

Technical field [0001] The present invention belongs to the field of biomedical detection, and more particularly to a cellular signal transduction activity detection method based on tetrahedral DNA nanomaterials. Background technique [0002] Cell signal transduction refers to cellular reactions of a series of effective gene expression through cellular factors and cell receptors, induced by biochemical reactions and interactions between intracellular proteins to induce a series of effective gene expression. Cell signal transduction participates in the main physiological functions such as proliferation, apoptosis, differentiation and metabolism. Cell signal transduction research plays a key role in revealing cell life activities and how various diseases occur. The transcription factor belongs to a special type of protein, which can be combined with a particular chromosome DNA sequence to regulate the expression of the gene. The activation of the transcription factor can represent ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6818G01N33/53G01N33/68G01N33/58
CPCC12Q1/6818G01N33/53G01N33/68G01N33/582G01N2333/4703C12Q2537/1373C12Q2563/107C12Q2525/151C12Q2525/161
Inventor 黄菲张莹
Owner THE FIRST AFFILIATED HOSPITAL OF FUJIAN MEDICAL UNIV
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