Detection method for expression of GPR120 gene based on eGFP, and application of eGFP to detection of expression of GPR120 gene

A detection method and gene expression level technology, applied in the field of eGFP biological applications, can solve the problems of cumbersome experimental steps and inability to monitor gene expression changes, etc., achieve simple and fast detection, reliable measurement results, and reduce the effect of group error

Inactive Publication Date: 2017-10-03
XIAN MEDICAL UNIV
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Problems solved by technology

[0004] The purpose of the present invention is to provide a method for detecting the expression level of GPR120 gene based on eGFP and its application, which solves the problems that the existing GPR120 gene expression level detection has complicated experimental steps and cannot monitor the change of gene expression at the living cell level. Provides a new idea for the detection of GPR120 gene expression level

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  • Detection method for expression of GPR120 gene based on eGFP, and application of eGFP to detection of expression of GPR120 gene
  • Detection method for expression of GPR120 gene based on eGFP, and application of eGFP to detection of expression of GPR120 gene
  • Detection method for expression of GPR120 gene based on eGFP, and application of eGFP to detection of expression of GPR120 gene

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Embodiment

[0050] The alveolar macrophages of eGFP-labeled GPR120 model mice were cultivated, and the specific method was as follows: in vitro culture, the eGFP-labeled GPR120 model mice were sacrificed by cervical dislocation, and the mice were sterilized with 75% alcohol; Draw about 6ml of PBS buffer solution, remove air bubbles for later use; use ophthalmic scissors to cut open the neck to the outer skin of the thoracic cavity, separate the trachea in the neck for threading, cut open the thorax to expose the lungs, clamp the front end of the trachea with tweezers, cut the trachea short and put the syringe Insert the needle tip into the trachea and tie it tightly with a thread; slowly inject 2-3ml of PBS buffer solution to fill the lungs, suck out the solution in the lungs and inject it slowly, cycle 3-5 times, absorb the PBS buffer solution in the lungs and inject it into the centrifuge tube, and the obtained lung giant Phage cell solution; balance the centrifuge tube and centrifuge in...

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Abstract

The invention discloses a detection method for the expression of a GPR120 gene based on eGFP. The detection method comprises the following steps: inserting an eGFP fragment to the specific site TAA of the termination codon of the GPR120 gene by using CRISPR/Cas 9 technology so as to obtain a transgenic model mouse with eGFP-labeled GPR120 positive cells; and carrying out excitation by using a fluorescence analyzer and determining the fluorescence intensity of the positive cells of the mouse. The method carries out real-time monitoring on the expression of the GPR120 gene so as to control and reduce errors among groups, guarantee the reliability of results and compensate for and overcome the problems of variability caused by comparison of different groups of histocytes and incapability of monitoring the changes of gene expression at a living cell level in the prior art. According to the invention, the expression level of the GPR120 gene can be immediately determined after collection of cells, and operations and reactions like RNA extraction, inverse transcription and PCR are omitted, so detection of the expression of the GPR120 gene is simpler.

Description

technical field [0001] The invention belongs to the technical field of eGFP biological application, and in particular relates to an eGFP-based detection method for GPR120 gene expression, and also relates to an eGFP-based application in the detection of GPR120 gene expression. Background technique [0002] GPR120 (G-protein coupled receptor 120; also known as free fatty acid receptor 4, FFAR4) is a member of the fatty acid receptor family and belongs to G protein coupled receptors (GPCR), which can be activated by long-chain fatty acids, especially n-3 Unsaturated fatty acids have the strongest activation ability. GPR120 is expressed in many tissues such as the brain, pituitary, lung, tongue, gastrointestinal tract, and adipose tissue, and is mainly distributed in various endocrine cells, adipocytes, macrophages, osteoblasts and osteoclasts, and taste bud cells in the gastrointestinal tract. The activation of GPR120 can stimulate the secretion of gastrointestinal hormones s...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/85
CPCC12N15/85C12Q1/6897
Inventor 赵玉峰苏兴利徐曦闫爱丽谢荣魏兰兰刘英光陈镝梁向艳赵妍妍
Owner XIAN MEDICAL UNIV
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