H3 subtype flu quick-detecting type classifying method based on RT-LAMP technology

Abstract

The invention discloses a H3 subtype flu quick-detecting type classifying method based on an RT-LAMP technique. The method comprises the following steps: firstly, analyzing the HA gene sequence of the H3 subtype flu virus in a flu virus gene group sequence database, designing highly special and conserved RT-LAMP special primers and six primers corresponding to eight areas of target genes, wherein four primers are FIP, BIP, F3 and B3, and the other two primers are loop primers LP1 and LP2; secondly, treating a sample to be detected in advance, and extracting the total RNA of the sample to be used as a reaction template; thirdly, configuring an RT-LAMP reaction system; fourthly, mixing the RNA template into the RT-LAMP reaction system, arranging the RT-LAMP reaction system in a water bath, and carrying out RT-LAM enlargement at constant temperature; and fifthly, identifying the enlarged reaction product. The result shows that the reaction product contains H3 subtype flu virus when the reaction product is positive; and the reaction product is H3 subtype flu virus when the reaction product is negative.

Description

Contents of the invention [0001] The invention relates to a method for rapid detection and typing of H3 subtype influenza based on RT-LAMP technology. Background technique [0002] Influenza, referred to as influenza, is an acute respiratory infectious disease caused by influenza virus. The incubation period is 1-3 days. The clinical manifestations are sudden onset, high fever, chills, headache, myalgia, general malaise. Generally, patients can recover in about a week, but for some special groups, such as infants and young children under 2 years old, elderly people over 65 years old, patients with underlying diseases or immunosuppressed patients, severe influenza and complications may occur in these groups The probability of death increases, and the fatality rate also increases accordingly. For a long time, influenza has been a serious infectious disease that endangers human health and public safety, and the annual economic losses caused by it are also very huge. Accordin...

AI Technical Summary

Problems solved by technology

Serological testing needs to collect double serum samples of patients in the acute phase and recovery phase, which is a retrospective diagnosis and has epidemiological significance, but it is of little significance for early acute diagnosis and has low sensitivity

Direct antigen detection methods include direct or indirect fluorescence test, enzyme-linked immunoassay, etc. The time is 2 hours to 1 day, which is suitable for early rapid diagnosis, but the sensitivity is low, and large sample detection cannot be performed

Molecular biology methods are a series of test methods for the detection of pathogenic nucleic acid molecules, mainly including RT-PCR, multiplex PCR, real-time PCR, NASBA, etc. The virus is quantified and typed, but the disadvantage is that it cannot perform simultaneous parallel detection and screening of a large number of clinical samples

[0005] In terms of sensitivity, specificity, rapidity, detection of a large number of specimens, and virus typing, all the above-mentioned detection methods have certain defects, and their application in clinical diagnosis and epidemiological monitoring is limited.

RT-PCR and fluorescence quantitative RT-PCR detection techniques have been widely used, and the results are reliable, but the detection time of fluorescence quantitative RT-PCR is short, but it requires expensive detection equipment, which is difficult to use in front-line detection units. Conventional RT-PCR Although the cost of the required PCR instrument is not as high as that of the quantitative RT-PCR instrument, the cost is also considerable, and it takes 2-3 hours to produce the result

The relevant instruments required for gene chip detection are more expensive and require professional technicians

Images