Detection method for EV71virus in environmental water body

A detection method, EV71-R technology, is applied in the field of environmental water quality detection, which can solve the problems of low EV71 concentration, no EV71 detection method, and inability to accurately qualitatively or quantitatively detect EV71.

CN103305628BInactive Publication Date: 2015-07-15XI'AN UNIVERSITY OF ARCHITECTURE AND TECHNOLOGY
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
Publication Date
2015-07-15
Estimated Expiration
Not applicable · inactive patent

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Abstract

The invention discloses a detection method for EV71 virus in an environmental water body, wherein a semi-nested RT-PCR (Reverse Transcription-Polymerase Chain Reaction) and real-time fluorescence quantification RT-PCR reaction system is established to detect EV71 virus in the environmental water body by using a recombinant plasmid as a reference product, and the recombinant plasmid is prepared by using a primer and a probe aiming at EV71 virus designed according to a genetic conservative fragment of enterovirus belonging to a VP1 serotype area. The method has good specificity and high sensitivity and precision, and can qualitatively and quantitatively detect EV71 virus in the environmental water body accurately.
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Description

technical field

[0001] The invention belongs to the technical field of detection of environmental water quality, and in particular relates to a detection method of EV71 virus in environmental water. technical background

[0002] Enterovirus 71 (EV71) belongs to the Enterovirus genus and has attracted much attention in recent years. It is a major pathogen that causes hand, foot and mouth disease. EV71 is a water-borne virus that can be leaked from the secretions of EV71-infected patients into environmental water bodies and can survive in environmental water bodies for a long time. The water body polluted by it becomes an important pollution source of EV71, which can cause the outbreak of hand, foot and mouth disease.

[0003] Existing methods for detecting EV71 mainly include cell culture method and polymerase chain reaction (Polymerase Chain Reaction, PCR). There are a variety of pathogenic microorganisms in environmental water, especially other enteroviruses. These microo...

Examples

Embodiment

[0075] The equipment and reagents used in this embodiment are as follows:

[0076] (1) High-speed refrigerated centrifuge with a maximum speed of 14000r / min;

[0077] (2) 4℃, -80℃ refrigerator

[0078] (3) Centrifuge tubes, 50mL, 1.5mL, 0.5mL, 0.2mL;

[0079] (4) Trace ultraviolet spectrophotometer;

[0080] (5) Constant temperature metal heater

[0081] (6) Bacterial Shaker, Bacterial Incubator

[0082] (7) Real-time fluorescent quantitative PCR instrument;

[0083] (8) gradient qualitative PCR instrument;

[0084] (9) Electrophoresis apparatus;

[0085] (10) Gel Imager

[0086] (11) pMD19-T vector;

[0087] (12) Competent Escherichia coli DH5α:

[0088] (13) Hydrochloric acid, NaCl solution, MgCl 2 solution, etc.;

[0089] (14) Polyethylene glycol (PEG-6000) with a molecular weight of 6000, glucose, etc.;

[0090] (15) 5mol / L NaCl solution;

[0091] (16) IPTG / X-gal / Amp plate;

[0092] (17) LB liquid medium;

[0093] (20) total RNA extraction kit;

[0094] (18...