Fluorescent quantitative RT-PCR detection reagent, kit and detection method for West Nile virus

Abstract

The invention discloses a fluorescent quantitation RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection reagent of a West Nile virus, a kit and a detection method of the West Nile virus. The detection reagent takes a conserved fragment of a West Nile virus E gene as a target, and comprises a pair of specific primers, a specific probe and an internal labeling probe. The kit comprises PCR mixed liquor, Taq DNA (Deoxyribonucleic Acid) polymerase, DEPC-H2O, a positive quality control, a negative quality control, a quantification standard and a pseudovirus internal labeling solution, wherein the PCR mixed liquor comprises a pair of specific primers, a specific probe and an internal labeling probe. The detection method comprises extraction of total RNAs (Ribonucleic Acids), preparation of reaction components, dilution of the standard for making a standard curve, amplification and result judgment. The reagent and the kit are high in specificity, sensitivity and flux and environmental friendly, and can be used for accurately detecting low-content West Nile virus infection, unapparent infection or continuous toxic hosts. The detection method is quick, specific, sensitive and quantitative, and can be used for detecting a large number of samples at the same time.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a PCR detection reagent, a kit and a detection method thereof, in particular to a fluorescent quantitative RT-PCR detection reagent, a kit and a detection method of West Nile virus. Background technique [0002] West Nile virus disease is a highly infectious zoonotic disease that causes serious public health problems. West Nile virus can be transmitted through blood vectors such as mosquitoes or ticks, and most of them lack effective treatment and preventive measures, which poses a severe challenge to disease control. There are many methods for serological diagnosis of West Nile virus, but these methods have certain limitations. For example, ELISA is not a specific diagnostic method and can only be used for screening of virus antibodies, but not for diagnosis; the specific plaque reduction neutralization test is complicated, time-consuming and laborious, and is difficult to apply to l...

AI Technical Summary

Problems solved by technology

For example, ELISA is not a specific diagnostic method, it can only be used for the screening of virus antibodies, but not for diagnosis; the specific plaque reduction neutralization test is complicated, time-consuming and laborious, and it is difficult to apply to large-scale censuses

The detection method of West Nile virus nucleic acid mainly includes common RT-PCR method, but its sensitivity is poor when detecting horse brain tissue disease materials

This method also has a big disadvantage: it is easy to pollute and prone to false positives

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