Primer group, kit and method for detecting listeria monocytogenes on basis of intelligent isothermal amplification technology

Abstract

The invention discloses a primer group, a kit and a method for detecting listeria monocytogenes on the basis of an intelligent isothermal amplification technology. The detecting primer group comprisesa TP primer, an FP primer, a BP primer, an OP1 primer and an OP2 primer. The detecting kit comprises a primer solution, a reaction solution, Bst DNA polymerase, mispairing binding protein, a color developing agent and controls. According to the detecting method, to-be-detected DNA is extracted, DNA polymerase activity with strand displacement activity is used, four specific primers and mispairingbinding protein are used, a sample DNA template is amplified at 60-65 DEG C, pg class of DNA can be detected, and the identification manner is performed as follows: SYBR(R) green I is added to a reaction system, whether amplification is performed is detected and judged by an ESE-tube scanner instrument, and whether the to-be-detected sample contains listeria monocytogenes DNA is determined. The method has the advantages of being rapid, efficient, convenient to operate, high in specificity, high in sensitivity, convenient in identification, suitable for field detection and the like and is suitable for promotion and application.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and relates to a primer set, a kit and a method for detecting Listeria monocytogenes based on an intelligent constant temperature amplification technology. Background technique [0002] Listeria monocytogenes (Listeria monocytogenes) is one of the key regulatory targets of the food safety department. It pollutes meat, milk, eggs, aquatic products and vegetables to varying degrees, and the pollution of meat and meat products is the most serious. Therefore, it is particularly important to establish a rapid, accurate, quantitative and reliable detection method for Listeria monocytogenes in meat. The current detection methods for common pathogenic bacteria in meat products are cumbersome, time-consuming, laborious, low sensitivity, poor specificity, and prone to false positives, which cannot meet the development needs of rapid identification of Listeria monocytogenes. With the development ...

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Problems solved by technology

The current detection methods for common pathogenic bacteria in meat products are cumbersome, time-consuming, laborious, low sensitivity, poor specificity, and prone to false positives, which cannot meet the development needs of rapid identification of Listeria monocytogenes

With the development of molecular biology, many identification methods based on molecular biology techniques have emerged, such as PCR-SSCP, PCR-RFLP, DNA probes, DNA chips, DNA direct sequencing, etc., with their advantages of high efficiency and specificity It is favored, but requires high technical level of testing personnel, expensive equipment, and long testing cycle. In order to meet the needs of rapid identification and classification, it provides a fast and convenient method for scientific research, shortens the identification time of Listeria monocytogenes, and reduces the identification time. Technical difficulty and cost have always been hot spots in related research

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