Recombination method and application of luciferase reporter gene with aquaporin 1 (AQP1) promoter to drug screening

A technology of aquaporin and reporter gene, which is applied in the field of constructing AQP1 promoter salivary gland cell model and drug screening, and can solve the problems that there are no therapeutic drugs.

Inactive Publication Date: 2013-08-07
JILIN UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

So far, there is no effective treatment drug, so it is of great practical significance to develop an effective treatment drug for Sjögren syndrome

Method used

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  • Recombination method and application of luciferase reporter gene with aquaporin 1 (AQP1) promoter to drug screening
  • Recombination method and application of luciferase reporter gene with aquaporin 1 (AQP1) promoter to drug screening
  • Recombination method and application of luciferase reporter gene with aquaporin 1 (AQP1) promoter to drug screening

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Experimental program
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Embodiment 2

[0068] Example 2: Drug screening

[0069] 1. Cultivation of cell lines

[0070] Adenoid cystic cancer cells were cultured in DMEM, supplemented with 10% fetal bovine serum, 100U / mL penicillin and 100ug / mL streptomycin. The cells were cultured in a 5% incubator at 37°C.

[0071] 2. Prove the effectiveness of the drug screening model

[0072] Scholars at home and abroad have confirmed that estradiol (E 2 ) Plays an important role in the gene expression of AQP1. The present invention uses AQP1 promoter luciferase reporter gene recombinant plasmid cell model to confirm that E 2 It can activate the AQP1 promoter in a dose-dependent manner [ Figure 8 】At the same time, we also confirmed that estradiol can promote the expression of AQP1 protein【 Picture 9 ], which proves that the cell model can be used for drug screening simply and sensitively.

[0073] 3. Application of drug screening

[0074] First take 2×10 5 ACC-M cells are plated in a 48-well plate to make the cells reach a confluence r...

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Abstract

The invention belongs to the technical field of bioengineering and in particular relates to a recombination method and application of a luciferase reporter gene with a human aquaporin 1 (AQP1) promoter to drug screening. The highly expressed AQP1 plays an important role in the salivary secretion process and is closely related to onset of Sjogren syndromes. In the invention, a 1868bp AQP1 promoter sequence is amplified from a personal genome by utilizing the gene engineering technology and is cloned onto a PGL2-luc vector to construct an AQP1 promoter-Luc expression vector. AQP1 expression is controlled by a promoter region. Constructing a salivary gland cell model with the AQP1 promoter is a key step for screening the traditional Chinese medicine ingredients which can treat the Sjogren syndromes. The drug screening cell model proves that ginseng and American ginseng ingredients at certain concentrations can promote transcription of the AQP1 promoter and the higher the concentrations are, the stronger the activity is, and can guide the drugs to be clinically applied to treat the Sjogren syndromes.

Description

Technical field [0001] The invention belongs to the technical field of bioengineering, and specifically relates to the recombination of the human aquaporin 1 (AQP1) promoter luciferase reporter gene, the construction of the AQP1 promoter salivary gland cell model and the application of drug screening. Background technique [0002] Sjogren’s syndrome is an autoimmune disease that mainly invades exocrine glands such as salivary glands and lacrimal glands. The main clinical manifestations are reduced saliva secretion, leading to dry lips, chapped, rampant caries, oral mucosal ulcers, fungal and viral infections accompanied by burning pain, foreign body sensation, etc. With the changes in people's lifestyles, the incidence rate is getting higher and higher, with an adult incidence rate of 0.5%-3.0%, and it is more common in middle-aged and elderly women, with a male to female ratio of 1:9. Up to now, there is no effective treatment drug, so the development of effective Sjogren syndr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/66C12N15/63C12Q1/68C12Q1/04
Inventor 李江赵超悦焦晓菲姜春娃贺玺魏溦张艳
Owner JILIN UNIV
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