Fluorescent quantitative RT-PCR detection kit and detection method for enterovirus

A detection kit, RT-PCR technology, used in fluorescence/phosphorescence, microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of inability to identify antigenic variants or new types, time-consuming and cumbersome

Inactive Publication Date: 2009-04-15
ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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Problems solved by technology

With the eradication of poliovirus, some new infectious diseases caused by enteroviruses are also popular from time to time, so the laboratory diagnosis of enteroviruses is becoming more and ...

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  • Fluorescent quantitative RT-PCR detection kit and detection method for enterovirus
  • Fluorescent quantitative RT-PCR detection kit and detection method for enterovirus
  • Fluorescent quantitative RT-PCR detection kit and detection method for enterovirus

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Embodiment 1

[0043] 1 Materials and methods

[0044]1.1 Virus strains and clinical specimens:

[0045] Enterovirus EV71, Coxsackie group A (CA) CA4, CA16, CA21, Coxsackie B5, E6, E30, POLIO type 1, POLIO type 3 and other virus strains were obtained from Chinese Academy of Medical Sciences, Beijing Institute of Biological Products and isolates from Zhejiang Provincial Center for Disease Control and Prevention. The clinical samples come from the cerebrospinal fluid, herpes fluid, feces, etc. of patients suspected of the recent enterovirus outbreak in Zhejiang Province. After the samples are collected, they are transported to the laboratory with ice.

[0046] 1.2 Primers and probes

[0047] Enterovirus strains from all over the world were downloaded from the NCBI gene bank in the United States, including all 67 serotypes of enteroviruses. The homology comparison was carried out, and primers and Taqman probes were designed in the non-coding general conserved gene region at the 5′ end of all...

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Abstract

The invention provides a fluorescence quantitative RT-PCR detecting kit for an enterovirus and a detecting method thereof; and the sequences of an upstream primer and a downstream primer and a specific probe of the fluorescence quantitative RT-PCR detecting kit are as follows: the upstream primer EV(YG)F is 5'-GGCTGCGYTGGCGGCC-3', the downstream primer EV(YG)R is 5'-CCAAAGTAGT CGGTTCCGC-3' and the specific probe EV(YG)PB is 5'-CTCCGGCCCCTGAATGCGG-3'. The method has high specificity on detecting the enterovirus and does not have cross reactions with other enteroviruses such as hepatitis A, hives, rubella, parotitis, encephalitis, dengue fever, adenovirus, and the like. The detection sensitivity of the method achieves 0.1TCID50; the method can directly detect the nucleic acid of the enterovirus from the samples of ncurolymph, herpes liquid, dejecta and the like of suspected patients; only about 3h is needed from extracting the nucleic acid of the enterovirus to finishing the detection; and the detecting kit and the detecting method are suitable for the lab early diagnosis of sudden epidemic caused by the infection of the enteroviruses such as the hand-foot-and-mouth disease and the like.

Description

(1) Technical field [0001] The invention relates to an enterovirus nucleic acid fluorescence quantitative RT-PCR detection kit and a detection method, which can be applied to laboratory emergency detection of outbreaks caused by enteroviruses. (2) Background technology [0002] Enteroviruses (EVs) belong to the picornavirus family and can be divided into polioviruses, coxsackieviruses, echoviruses and enteroviruses 68-71 according to serology, with a total of 67 serotypes. Enteroviruses can cause many diseases, and can break out and cause death, seriously endangering human health. With the eradication of poliovirus, some new infectious diseases caused by enteroviruses are also popular from time to time, so the laboratory diagnosis of enteroviruses is becoming more and more important, but the traditional serum neutralization test is not only tedious, time-consuming, but also It is impossible to identify all serotypes, let alone some antigen variants or new types. The fluore...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68G01N21/64
CPCY02A50/30
Inventor 卢亦愚严菊英徐昌平冯燕陈寅
Owner ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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