Multiple rapid detection method, detection primer group and kit for three food-borne pathogenic bacteria

A rapid technology for detection of primers, applied in the field of microbial detection, can solve problems such as inability to identify multiple pathogenic bacteria, and achieve the effects of optimization of the LAMP reaction system, strong specificity, and optimized reaction conditions

Inactive Publication Date: 2012-01-25
浙江省质量技术监督检测研究院
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Problems solved by technology

[0006] The purpose of the present invention is to provide a multiple rapid detection method for three kinds of food-borne pathogenic bacteria and detection primer set and kit, so as to overcome the current existing methods that can only detect a certain pathogenic bacteria in one system , but cannot simultaneously judge whether there are multiple pathogenic bacteria in the same system

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  • Multiple rapid detection method, detection primer group and kit for three food-borne pathogenic bacteria

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Embodiment 1

[0029] Embodiment 1: Optimum Mg in the LAMP reaction system of triple LAMP rapid detection method of the present invention 2+ Determination of concentration.

[0030] The specific determination method of this embodiment is:

[0031] 1. Extraction of sample DNA

[0032] 11 Inoculate the standard strains of Salmonella and Staphylococcus aureus in nutrient broth respectively, inoculate Listeria monocytogenes in brain heart infusion broth, and incubate at 36°C±1°C for 18 hours.

[0033] 1.2 Use the QIAGEN Bacterial Genomic DNA Extraction Kit to extract the bacterial DNA in the above cultured products respectively.

[0034] 2. LAMP reaction

[0035] 2.1 Artificially synthesized primer sets for the detection of Salmonella, Staphylococcus aureus and Listeria monocytogenes respectively. Wherein, the upstream primer of the outer primer used in the primer set for Salmonella detection has a sequence as shown in SEQ No.1, the downstream primer of the outer primer has a sequence as sho...

Embodiment 2

[0050] Example 2: Determination of the optimal LAMP reaction temperature of the triple LAMP rapid detection method of the present invention.

[0051] The specific determination steps of this embodiment are as follows:

[0052] 1. Extraction of sample DNA

[0053] 1.1 Inoculate the standard strains of Salmonella and Staphylococcus aureus in the nutrient broth respectively, inoculate the Listeria monocytogenes in the brain heart infusion broth, and incubate at 36°C±1°C for 18 hours.

[0054] 1.2 Use the QIAGEN Bacterial Genomic DNA Extraction Kit to extract the bacterial DNA in the above cultured products respectively.

[0055] 2. LAMP reaction

[0056] 2.1 Artificially synthesize primer sets for the detection of Salmonella, Staphylococcus aureus and Listeria monocytogenes respectively, and the primers and sequences of each primer set are consistent with those described in Section 2.1 of Example 1.

[0057] 2.2 LAMP reaction system

[0058] 2.2.1 LAMP reaction system of Salm...

Embodiment 3

[0071] Example 3: The detection sensitivity of the triple LAMP rapid detection method of the present invention to Salmonella.

[0072] The concrete detection method of the present embodiment is:

[0073] 1. Extraction of sample DNA

[0074] 1.1 Inoculate the standard strain of Salmonella in nutrient broth and incubate at 36°C±1°C for 18 hours.

[0075] 1.2 Use the QIAGEN Bacterial Genomic DNA Extraction Kit to extract the bacterial DNA in the above cultured products.

[0076] 2. LAMP reaction

[0077] 2.1 Artificially synthesize primer sets for the detection of Salmonella, Staphylococcus aureus and Listeria monocytogenes respectively, and the primers and sequences of each primer set are consistent with those described in Section 2.1 of Example 1.

[0078] 2.2 Reaction system of LAMP

[0079]Set up 9 reaction tubes, the total volume of the reaction system in each reaction tube is 25 μL, including: the upstream primer and downstream primer of the external primers for the det...

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Abstract

The invention discloses a multiple rapid detection method, a detection primer group and a kit for three food-borne pathogenic bacteria. According to the invention, upstream and downstream primers of an external primer and upstream and downstream primers of an internal primer of a rapid detection primer group of salmonella correspondingly have sequences as shown in SEQNo.1, SEQNo.2, SEQNo.3 and SEQNo.4; upstream and downstream primers of an external primer and upstream and downstream primers of an internal primer of a rapid detection primer group of staphylococcus aureus have sequences as shown in SEQNo.5, SEQNo.6, SEQNo.7 and SEQNo.8; upstream and downstream primers of an external primer and upstream and downstream primers of an internal primer of a rapid detection primer group of Listeria monocytogenes have sequences as shown in SEQNo.9, SEQNo.10, SEQNo.11 and SEQNo.12. By using the method in the invention, the defect that the prior art can not be used for simultaneously judging whether multiple pathogenic bacteria exist in the same system at the same time is overcome, and whether the same detected sample contains one or more of salmonella, staphylococcus aureus and Listeria monocytogenes can be correctly detected.

Description

technical field [0001] The invention belongs to the field of microorganism detection, and relates to a triple rapid detection method for Salmonella, Staphylococcus aureus and Listeria monocytogenes, a detection primer set and a detection kit. Background technique [0002] Food safety is a major concern of all countries in the world, and food-borne pathogens are one of the main reasons affecting food safety. Salmonella (Salmonella spp) is an important pathogenic bacterium in the field of public health. According to statistics, in the bacterial food poisoning that occurs in various countries in the world, the food poisoning caused by Salmonella often ranks first, and the bacterial food poisoning that occurs in the inland areas of my country is also Salmonella was the leading cause. In addition to causing food poisoning, bacteria in this genus can also cause diseases such as gastroenteritis, typhoid fever, and paratyphoid fever. Staphylococcus aureus (Staphylococcus aureus) be...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68C12Q1/04C12Q1/14
CPCY02A50/30
Inventor 姜侃张东雷金燕飞汪新黄建锋陈小珍
Owner 浙江省质量技术监督检测研究院
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