LAMP detection reagent kit for RNA virus, detection method and application

Abstract

The invention discloses a loop-mediated isothermal amplification (LAMP) detection reagent kit for the RNA virus. The LAMP detection reagent kit comprises a hydration solution A, a hydration solution B, a positive control, mineral oil and a negative control. The hydration solution A is prepared from Tris-HCl, KCl, MgCl2, Triton X-100, reverse transcriptase, dNTPs, an RNAase inhibitor, a random primer, sodium dodecyl sulfate, polyoxyethylene octylphenol ether (10) sodium succinate, beta-mercaptoethanol, EDTA, glycerol, dithiothreitol, PEG-4000 and the balance double distilled water. The hydration solution B is prepared from Tris-HCl, KCl, MgSO4, (NH4)2SO4, dNTPs, an inner primer FIP, an inner primer BIP, an outer primer F3, an outer primer B3, Bst DNA polymerase, beta-mercaptoethanol, EDTA, glycerol, dithiothreitol, tetramethyl benzidine, H2O2 and a virus specific recognition sequence. The reagent kit can efficiently, quickly, accurately and sensitively obtain a detection result of the virus, the detection result is high in specificity and sensitivity and can be easily judged with the naked eyes, and the detection reagent kit is suitable for field quick detection.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a LAMP detection kit, detection method and application of an RNA virus. Background technique [0002] At present, the traditional methods for detecting and diagnosing viral diseases include virus neutralization test, enzyme-linked immunosorbent assay, immunofluorescence antibody test, immunofluorescence electron microscopy and PCR, etc. These methods have been used in the detection of pathogens and the diagnosis and research of diseases. It has played a huge role, but there are problems such as time-consuming, complicated operation, expensive equipment and instruments, and it is not conducive to on-site detection, which limits their application in rapid virus diagnosis. [0003] Loop-mediated isothermal gene amplification (LAMP) is a new type of gene amplification method, which has the following characteristics: simple, fast, does not require expensive equipmen...

AI Technical Summary

Problems solved by technology

[0002] At present, the traditional methods for detecting and diagnosing viral diseases include virus neutralization test, enzyme-linked immunosorbent assay, immunofluorescence antibody test, immunofluorescence electron microscopy and PCR, etc. These methods have been used in the detection of pathogens and the diagnosis and research of diseases. It has played a huge role, but the problems are that it takes a long time, the operation is complicated, expensive equipment and instruments are required, and it is not conducive to on-site detection, etc., which limits their application in rapid virus diagnosis

The first two judgment methods have the disadvantages of complex operation and need special equipment. The method of adding color developer to observe the color is easy to operate and does not require special equipment.

At present, there are two kinds of chromogenic reagents used in LAMP: DNA intercalation dye and metal ion indicator, but the specificity and sensitivity of these two chromogenic reagents are not high, and the results are not easy to judge by naked eyes, which limits their application. Therefore, In order to obtain virus detection results efficiently, quickly, accurately and sensitively, it is necessary to develop a new detection kit containing a color reagent with high specificity, strong sensitivity and easy judgment by naked eyes.

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