Visual detection method of isothermal amplification products of nucleic acids

Abstract

The present invention relates to two visual detection reagents of isothermal amplification products of nucleic acids, the two visual detection reagents are that 1-(1-hydroxy-2-naphthalene azo)-6-nitro-2-naphthol-4-sodium sulfonate or derivatives thereof (hereinafter referred to as the reagent 1) and 4-(2-pyridine azo) resorcinol sodium salt or derivatives thereof (hereinafter referred to as the reagent 2), the two visual detection reagents are used alone, at the beginning of a reaction, the reagent 1 combines with magnesium ions to become red, and the reagent 2 combines with the magnesium ions to form an orange complex, with the reaction, by-product pyrophosphate groups combines the magnesium ions to form a precipitation, the reagent 1 or loses the magnesium ions to become blue from red, the reagent 2 becomes yellow from orange, amplification reaction results can be judged by color change, detection without opening of a cover can be realized by the method, the aerosol pollution can be overcome, and the false positive problem caused by primer dimmers can be solved.

Description

technical field [0001] The invention relates to two types of nucleic acid isothermal amplification product visualization detection reagent 1-(1-hydroxy-2-naphthazo)-6-nitro-2-naphthol-4-sodium sulfonate or its derivatives (hereinafter referred to as reagents) 1) or 4-(2-pyridylazo)resorcinol sodium salt or its derivatives (hereinafter referred to as reagent 2). Background technique [0002] With the development of modern molecular biology and molecular technology, many nucleic acid amplification techniques have been researched and developed. Among them, the nucleic acid isothermal amplification technology, especially the nucleic acid loop-mediated isothermal amplification technology (loop-mediated isothermal amplification, LAMP), due to its strong specificity, high sensitivity, and fast reaction speed, has been used in nucleic acid-specific amplification and The detection field has a tendency to replace PCR and has become one of the hot spots in the field of life science re...

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Problems solved by technology

The electrophoresis method requires the use of highly carcinogenic fluorescent dyes such as ethidium bromide, which not only endangers the health of the operator, but also causes cross-contamination; because the turbidity produced by the amplification reaction is not obvious, the turbidity method is easily affected by the subjective factors of the test personnel. The fluorescent dye method is mainly based on the detection of the amplified product DNA, while the LAMP reaction requires many pairs of primers, and the generation of primer dimers is inevitable, so the problem of false positives cannot be ruled out; the immunological lateral flow test paper requires After the amplification reaction, opening the cover and loading the sample for chromatography cannot overcome the problem of aerosol pollution; the market price of a real-time turbidimeter exceeds 400,000, which is much higher than the price of a PCR instrument, which limits the promotion of this technology in basic testing departments and medical departments application

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