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Method for screening influenza virus resisting medicines

An influenza virus, active technology, applied in the field of drug screening, can solve the problem of high toxicity and side effects of host cells, and achieve the effect of solving the problem of drug resistance

Inactive Publication Date: 2011-05-11
MEDICINE & BIOENG INST OF CHINESE ACAD OF MEDICAL SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Since the currently used anti-influenza drugs are mainly aimed at gene targets with strong variability in virus particles, the virus can always quickly adapt and escape the selection pressure imposed by the drug, resulting in drug resistance. There is a problem of serious side effects

Method used

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  • Method for screening influenza virus resisting medicines
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Experimental program
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Effect test

Embodiment 1

[0018] 1. Construction of expression vectors;

[0019] The Rluc gene fragment in the eukaryotic cell expression vector pRluc-N2 (PerkinElmer) was amplified by PCR. Primers used: the upstream primer 5'-GATTCAATTCAGCTACCATAATGATGCAGAGAGGCAATTTTGGTAC-3', the downstream primer 5'-CAAAATTGCCTCTCTGCATCATTATGCTAGCTGAATTG-3'.

[0020] reaction system:

[0021]

[0022]

[0023] Reaction conditions: 94°C for 45s, 68°C for 90s, a total of 30 cycles.

[0024] The Rluc fragment was inserted into the cloning site BsmB I of the influenza virus M fragment expression vector pWSN-M (gifted by Dr. Kawaoka) to obtain the plasmid pFLU-Rluc, which expresses the fluorescent protein.

[0025] 2. Cell culture

[0026] 293T cells were cultured, and after the cells filled the culture flask, the old medium was discarded and digested with a digestive solution containing 0.25% trypsin and 0.02% EDTA. When the cells become round, discard the digestion solution, immediately add high-sugar DMEM med...

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Abstract

The invention discloses a method for screening influenza virus resisting medicines. In the method, a sample to be detected is added into a fusion protein cell which expresses influenza virus ribonucleic acid (RNA) polymerase and an activity-dependent reporter gene thereof, and the quantity of a coding product of the reporter gene is observed, so that the blocking effect of the sample on the RNA polymerase is determined; and whether the sample subjected to primary selection is an RNA polymerase activity inhibitor or not is further judged, so that the influenza virus resisting medicines are screened. By the screening method, an influenza virus RNA polymerase complex is taken as a target, so that the problem of medicine resistance caused by high virus variation is solved, and the cross medicine resistance of the screened influenza virus resisting medicines and the conventional clinically applied influenza virus resisting medicines is avoided.

Description

technical field [0001] The invention relates to a drug screening method, in particular to a screening method for anti-influenza virus drugs. Background technique [0002] Influenza (abbreviated as flu) is an acute respiratory infectious disease caused by influenza virus (influenza), mainly transmitted by air droplets. It is a worldwide infectious disease that humans cannot effectively control at present. It causes considerable morbidity and mortality every year, and influenza epidemics or local outbreaks occur from time to time. my country is recognized by the world as a frequent occurrence of influenza. [0003] At present, the means of clinically preventing and treating viral influenza mainly include drug therapy and vaccine. Because influenza virus is a multi-segment negative-strand RNA virus, it has strong recombination variation and antigenic drift characteristics, which seriously restricts the immune protection effect of current influenza vaccine and the therapeutic...

Claims

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Application Information

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IPC IPC(8): C12N5/10C12N15/85C12Q1/02G01N21/64
Inventor 岑山魏晓露刘振龙李晓宇张全贾平平杨亮
Owner MEDICINE & BIOENG INST OF CHINESE ACAD OF MEDICAL SCI
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