Detection method of coxsackievirus A10 in mixed infection of hand-foot-mouth disease

A technology of mixed infection and hand, foot and mouth disease, applied in the biological field, can solve the problems of huge cost and increase cost, and achieve the effect of reducing cost

Inactive Publication Date: 2018-03-20
INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This requires two RT-PCRs, which increases the cost, especially for large-scale molecular epidemiological investigations, which cost a lot. Therefore, it is necessary to explore a simple and fast CV-A10 detection method

Method used

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  • Detection method of coxsackievirus A10 in mixed infection of hand-foot-mouth disease
  • Detection method of coxsackievirus A10 in mixed infection of hand-foot-mouth disease
  • Detection method of coxsackievirus A10 in mixed infection of hand-foot-mouth disease

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Embodiment

[0020] 1. Sample processing

[0021] Take 0.5 g of feces, add 2.5 mL of PBS to shake and mix well, centrifuge at 3000 r / min for 30 min, take the supernatant, and store it at -20 °C;

[0022] 2. Viral RNA extraction

[0023] The extraction of viral RNA was performed according to the instructions of the kit (Qiamp viral RNA mini Kit), as follows: Take 140 μL of supernatant or cerebrospinal fluid and add 560 AVL, mix it upside down and place it at room temperature for 10 min; add 560 μL of absolute ethanol, mix it upside down; mix 630 μL of the above solution Add to the adsorption column, centrifuge at 8000rpm for 1min, add 500μLAW1, centrifuge at 8000rpm for 1min, pour off the waste liquid in the collection tube; add 500μLAW2, centrifuge at 14000rpm for 3 minutes, pour out the waste liquid in the collection tube; put the adsorption column Put it into an empty collection tube, centrifuge at 14000rpm for 1min; put the adsorption column into a new centrifuge tube, add elution buff...

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Abstract

The invention discloses a detection method of coxsackievirus A10 (CV-A10) in mixed infection of hand-foot-mouth disease. The method is as follows: extracting viral RNA, using enterovirus universal primer 224 and 222 for RT-PCR amplification, then using primers AN89 and AN88 for nested PCR amplification, if the virus is non-CV-A10 serotype after comparison, designing specific CV-A10 upstream primerCVA10TF and downstream primer CVA10TR according to the CV-A10 sequence, using the RT-PCR product as a template for nested PCR amplification, and taking the PCR product for 1% agarose gel electrophoresis to identify. The method has the advantages of simple operation, rapid detection, high accuracy, low cost, and the like, and is particularly suitable for large-scale molecular epidemiological investigation.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a detection method for Coxsackie A group 10 virus in clinical samples of hand, foot and mouth disease. Background technique [0002] Since 2008, in China, there have been more than one million cases of Hand-foot-mouth disease (HFMD) every year, and some deaths have been reported. HFMD has become one of the major public health problems in my country and Southeast Asian countries. HFMD is a common infectious disease caused by a variety of human enteroviruses. The pathogens causing HFMD are mainly CV-A16, 4, 5, 7, 9, 10, and B 3, 5, echovirus and EV-A71, among which Coxsackievirus A10 (CV-A10) has become one of the main pathogens besides enterovirus 71 (EV-A71), CV-A16 and CV-A6. [0003] During an outbreak of HFMD, there are multiple enterovirus co-infections, which may aggravate the disease and / or prolong the course of the disease. Therefore, the monitoring of mixed inf...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6848
CPCC12Q1/6848C12Q2531/113C12Q2549/119
Inventor 马绍辉张捷杨昭庆刘红波赵义林张海浩黄小琴孙浩
Owner INST OF MEDICAL BIOLOGY CHINESE ACAD OF MEDICAL SCI
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