TRPV4-His499 protein sensitive to magnetic force, magnetic regulation tool and application

A technology of trpv4-his499 and magnetic regulation, applied in the field of system neuroscience, can solve the problems of consuming large magnetic nanoparticles and heat, specific modification of magnetic nanoparticles, and inability to accurately target cells

Active Publication Date: 2019-04-19
ACADEMY OF MILITARY MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, in this experiment, the magnetic nanoparticles were not specifically modified, which has a high off-target effect and cannot accurately target the target cells, requiring a large amount of magnetic nanoparticles and heat

Method used

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  • TRPV4-His499 protein sensitive to magnetic force, magnetic regulation tool and application
  • TRPV4-His499 protein sensitive to magnetic force, magnetic regulation tool and application
  • TRPV4-His499 protein sensitive to magnetic force, magnetic regulation tool and application

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preparation example Construction

[0033] In the present invention, the preparation method of the functionalized modified magnetic nanoparticles preferably includes: encapsulating the carboxylated magnetic nanoparticles with an anti-His antibody.

[0034] In the present invention, there is no special limitation on the source of the carboxylated magnetic nanoparticles, and commercial products can be used. In the present invention, there is no special limitation on the encapsulation method of the anti-His antibody on the carboxylated magnetic nanoparticles, and conventional methods can be used.

[0035] In the present invention, the anti-His antibody on the functionally modified magnetic nanoparticles can specifically react with the His tag of the TRPV4-his499 protein, so that the magnetic induction protein is fixed and dispersed on the cell surface, and under the action of a magnetic field, the The effector specifically opens TRPV4 protein ion channels, triggering cellular calcium influx for neural calcium signa...

Embodiment 1

[0042] 1. Preparation of experimental reagents

[0043] The carboxylated magnetic nanoparticles of 1.20nm, referred to as MNP (with ferric oxide core) were purchased from Chemicell, and were coated with antibodies according to the standard experimental method provided by Chemicell ( figure 1 shown), to obtain magnetic nanoparticles (MNPs). That is, under the catalysis of compounds such as EDC and NHS, the carboxyl groups on the magnetic nanoparticles and the amino groups on the His antibody can react to form amide bonds to achieve stable bonding and achieve antibody encapsulation of magnetic nanoparticles.

[0044] 2. The TRPV4-Hi499 fragment was synthesized by gene synthesis, that is, a 6×his tag was inserted at the 499th amino acid position of the TRPV4 protein, and the fragment was connected to the backbone plasmid through a restriction site to obtain pcDNA3.1-TRPV4- His499-p2A-mcherry, this plasmid is used for HEK293T cell transfection.

[0045] 3. Molecularly construct ...

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Abstract

The invention provides a TRPV4-His499 protein sensitive to magnetic force, a magnetic regulation tool and application, and relates to the technical field of system nerve science. TRPV4-His499 proteinis capable of sensing magnetic fields and being activated with the aid of functionally modified magnetic nanoparticles, followed by triggering cellular calcium influx for neurocalcium signaling. The specific operation is as follows: inserting a 6*his label at the 499th position amino acid position of the TRPV4 protein to obtain the TRPV4-His499 protein sensitive to magnetic force, transferring theTRPV4-His499 protein into the brain of a mouse by using virus, simultaneously injecting magnetic nanoparticles wrapped by anti-His antibodies, and inducing cell calcium influx under the action of a magnetic field, can be used for conducting nerve calcium signal and regulating mouse behaviors.

Description

technical field [0001] The invention relates to the technical field of system neuroscience, in particular to a TRPV4-His499 protein sensitive to magnetic force, a magnetic regulation tool and application. Background technique [0002] At present, optogenetics-based neurostimulation technology has been widely developed and used in the deconstruction of most animal behavior-related neural mechanisms. As a new type of non-invasive deep brain stimulation tool, magnetogenetic technology has very important application potential in the field of nervous system research because of its advantages different from optogenetics. [0003] 1. Neural regulation and behavior research [0004] How the brain guides animal behavior is an important research goal in systems neuroscience research. Deeply stimulating the nervous system for artificial regulatory intervention is the main means of studying the functions of various brain regions and the interconnections between brain regions, and can ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/47C07K19/00C12N15/85
CPCC12N15/85C07K14/47C07K2319/21
Inventor 王常勇吴松芳周瑾李红
Owner ACADEMY OF MILITARY MEDICAL SCI
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