LAMP (loop-mediated isothermal amplification) detection kit for DNA (deoxyribonucleic acid) viruses, detection method and application

Abstract

The invention discloses an LAMP (loop-mediated isothermal amplification) detection kit for DNA (deoxyribonucleic acid) viruses. The LAMP detection kit comprises the following parts including a hydration solution, a positive control part, mineral oil and a negative control part, in addition, each liter of hydration solution is prepared from the following ingredients including 20mmol Tris HCl with the pH being 8.8, 10mmol of KCl, 6.5 mmol of MgSO4, 10mmpl of (NH4)2SO4, 1mL of Triton X-100, 0.2mmol of dNTPs, 1.6 mumol of inner primers FIP and 1.6 mumol of inner primers BIP, 0.2 mumol of outer primers F3, 0.2 mumol of outer primers B3, 480U of Bst DNA polymerase, 0.4 to 0.7g of lauryl sodium sulfate , 0.7 to 1.2g of polyoxyethylene octylphenol ether (10) ether sodium succinate, 5mmol of beta-mercaptoethanol, 6.7 mumol of EDTA, 100mL of glycerol, 30mmoL of dithiothreitol, 8 to 12g of PEG-4000, 0.1g of tetramethyl benzidine, 0.1mL of H2O2, 1.0 mumol of viral specific recognition sequences and the balance distilled water. The kit can be used for efficiently, fast, accurately and sensitively obtaining the virus detection result; in addition, the specificity of the detection result is high; the sensitivity is high; the judgment can be easily made by naked eyes; the detection kit is suitable for in-site fast detection.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a DNA virus LAMP detection kit, detection method and application. 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] Traditional polymerase chain reaction (PCR) is a technique for amplifying DNA sequences in vitro, and its basic principle is similar to the natural replication process of DNA. Since the esta...

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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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