LAMP (loop-mediated isothermal amplification) double-chain detection probe, kit and detection method for staphylococcus aureus

Abstract

The invention provides an LAMP (loop-mediated isothermal amplification) double-chain detection probe, a kit and a detection method for staphylococcus aureus, and belongs to the technical field of biology. The double-chain detection probe comprises a light-emitting probe and a quenching probe, the light-emitting probe comprises a conserved sequence and a loop primer LF which are connected in sequence, and the 5'end of the conserved sequence is marked with a fluorescent light-emitting group; conserved sequences of the quenching probe and the light-emitting probe are complementary, and the 3'end of the quenching probe is marked with a fluorescence quenching group. The LAMP detection kit comprises the double-chain detection probe, the kit and the detection method, the staphylococcus aureus isdD gene is used as a detection target, a double-chain probe-constant temperature color changing system is adopted, a result can be interpreted by combining fluorescent quantitative PCR map analysis and a naked eye observation method, the probe is high in specificity, the color changing color contrast is obvious, misjudgment is not prone to being generated, and the detection accuracy is high. The dual systems reflect each other, and the detection accuracy is high.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a LAMP double-stranded detection probe of Staphylococcus aureus, a kit and a detection method. Background technique [0002] Staphylococcus aureus is Gram-positive, is a spherical bacteria without spores, flagella, lactic acid fermentation, facultative anaerobic, is a common food-borne pathogenic microorganism, often parasitic on the skin, It is also ubiquitous in the nose, throat, stomach, sores, and the environment, often causing harm to the human body. [0003] At present, the commonly used detection methods for Staphylococcus aureus include chromogenic medium culture method, fluorescence quantitative PCR, LAMP constant temperature PCR method, etc., all of which can be effectively detected. However, the chromogenic culture method has disadvantages such as time-consuming and large human factors. It often takes a week to detect a sample, and there are also defects such ...

AI Technical Summary

Problems solved by technology

[0004] There are mainly 1 methods for interpreting the results of the ring-mediated constant temperature amplification method: turbidity method, which can be used to observe whether there is white flocculent precipitate with the naked eye after the reaction, and determine the test result. The subjective factors are strong and the sensitivity is low.

2. Chromogenic method, add nucleic acid dyes such as SYBR GREEN, HNB, calcein, etc. after the reaction, and display different colors under ultraviolet light or naked eyes. The result is easy to judge, but it needs to be opened after the reaction, which is very easy to cause aerosol Pollution

3. Fluorescence detection method. By adding fluorescent dye SYBR GREEN to the reaction system, the fluorescence changes in the system can be observed in real time through a fluorescent PCR instrument to generate a fluorescence curve, and the amplification results can be observed through the curve. This method is closed-tube detection to eliminate gas Sol pollution, but due to the characteristics of SYBR GREEN dye, it can combine with any double-stranded DNA and generate fluorescence, which is low in specificity and prone to false positives

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