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LAMP kit for rapid detection of Salmonella

A Salmonella and kit technology, applied in the field of LAMP kits, can solve the problems of low sensitivity, poor specificity, false positive application, etc., and achieve the effect of high repeatability, fast detection speed and high sensitivity

Inactive Publication Date: 2013-01-02
WUHAN ZHENFU PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The traditional culture method for detecting Salmonella requires separation, screening, biochemical identification, and serological identification if necessary, which generally takes 4 to 6 days, which is time-consuming and laborious, and has the disadvantages of low sensitivity, poor specificity, false positives, time-consuming and laborious, etc.
Various conventional PCR methods have the advantages of strong sensitivity, high specificity, simplicity, and rapidity. There are many reports on the detection of Salmonella, but there are problems such as false positives caused by contamination caused by post-PCR post-processing and the need for special equipment. which limits its application in grassroots units

Method used

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  • LAMP kit for rapid detection of Salmonella
  • LAMP kit for rapid detection of Salmonella
  • LAMP kit for rapid detection of Salmonella

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0098] 1. The composition and preparation of the new rapid visual detection of Salmonella LAMP kit, the composition of the reagents:

[0099] a) LAMP reaction mixture

[0100] The LAMP reaction mixture consists of LAMP 10×buffer 2.5μl, 10μmol / L inner primers FIP and BIP 2.0μl each, 10μmol / L outer primers F3 and B3 each 1.0μl, 10mmol / L dNTPs 3.0μl, 50mmol / LMgSO 4 3.0 μl, 4.5 μl of 5.0mol / L betaine, 1.0 μl of Bst DNA polymerase large fragment, and the total volume of the reaction solution is 20 μl.

[0101] The primer sequences are as follows:

[0102] Upstream outer primer F3: 5'-CGGCCCGATTTTCTCTGG-3';

[0103] Downstream outer primer B3: 5'-CGGCAATAGCGTCACCTT-3';

[0104] Upstream inner primer FIP: 5'-ATCCGCATCAATAATACCGGCCTTTGGTATGCCCGGTAAACAGA-3';

[0105] Downstream inner primer BIP: 5'-GAACGGCGAAGCGTACTGGACATCGCACCGTCAAAGGAA-3';

[0106] b) Standard positive template

[0107] The standard positive template pMD18-T-invA is a pMD18-T-invA vector containing a nucleotide...

Embodiment 2

[0117] 1. The composition and preparation of the novel rapid visual detection of Salmonella LAMP kit is different from the kit described in Example 1 in that the specific primer sequences are different. The primer sequences of this kit are as follows:

[0118] Upstream outer primer F3: 5'-CGGCCCGATTTTCTCTGG-3';

[0119] Downstream outer primer B3: 5'-CGGCAATAGCGTCACCTT-3';

[0120] Upstream inner primer FIP: 5'-GCGCAGCATCCGCATCAATAATGGTATGCCCGGTAAACAGAT-3';

[0121] Downstream inner primer BIP: 5′-GCGAACGGCGAAGCGTACTGTCGCACCGTCAAAGGAAC-3′;

[0122] 2. The method for detecting Salmonella using the described PCR kit based on the rapid detection of pathogenic Salmonella in industrial food based on the loop-mediated isothermal amplification technology is exactly the same as the method described in Example 1, repeating the test 3 times, and the obtained detection results are identical, There were precipitation in the experimental group, and no precipitation in the control group (...

Embodiment 3

[0124] 1. The composition and preparation of the novel rapid visual detection of Salmonella LAMP kit is different from the kit described in Example 1 in that the specific primer sequences are different. The primer sequences of this kit are as follows:

[0125] Upstream outer primer F3: 5'-CGGCCCGATTTTCTCTGG-3';

[0126] Downstream outer primer B3: 5'-CGGCAATAGCGTCACCTT-3';

[0127] Upstream inner primer FIP: 5'-GCGCAGCATCCGCATCAATAATATGCCCGGTAAACAGATGAG-3';

[0128] Downstream inner primer BIP: 5′-GCGAACGGCGAAGCGTACTGTCGCACCGTCAAAGGAAC-3′;

[0129] 2. The method for detecting Salmonella using the described PCR kit based on the rapid detection of pathogenic Salmonella in industrial food based on the loop-mediated isothermal amplification technology is exactly the same as the method described in Example 1, repeating the test 3 times, and the obtained detection results are identical, There were precipitation in the experimental group, and no precipitation in the control group (...

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Abstract

The invention discloses an LAMP (loop-mediated isothermal amplification) kit for rapid detection of Salmonella. The kit is composed of an LAMP reaction solution, a standard positive template, and a negative quality control standard. The LAMP reaction solution contains a Bst DNA polymerase large fragment, primers, an LAMP10*buffer, a dNTPs solution, an MgSO4 solution and betaine. The primers include forward and reverse primers. The kit disclosed in the invention has the advantages of good specificity, high sensitivity, rapidness and convenience, high repeatability, and judgeable results obtained by naked eyes, etc., can perform rapid qualitative detection on pathogenic Salmonella in industrial food, and can be used to replace the continuously used traditional culture method and the serological diagnostic method.

Description

technical field [0001] The invention relates to a LAMP kit for rapid detection of Salmonella, belonging to the field of food detection. Background technique [0002] Salmonella is one of the important pathogens causing bacterial food poisoning, and it is also the most complex genus of Enterobacteriaceae and is a Gram-negative bacteria. At present, 2523 serotypes have been isolated in the world, and 216 serotypes have been found in my country. Various serotypes of Salmonella can cause poisoning, the most common being Salmonella enteritidis, Salmonella typhimurium and Salmonella choleraesuis. The symptoms of salmonella poisoning are mainly acute gastroenteritis. The incubation period is generally four to forty-eight hours, the short-term is several hours, and the long-term is two to three days. The early symptoms include nausea, headache, malaise and chills, etc. The main symptoms are vomiting , Diarrhea, abdominal pain, yellow-green watery stool, sometimes with pus, blood a...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12R1/42
Inventor 王业富郑虎卢晅
Owner WUHAN ZHENFU PHARMA CO LTD
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