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Salmonella enteritidis detection reagent kit and method based on loop-mediated isothermal amplification technology

A Salmonella Enteritidis, ring-mediated isothermal technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems that have not been seen in the detection of Salmonella Enteritidis, and achieve simple and fast operation and low detection cost The effect of low and high sensitivity

Inactive Publication Date: 2013-01-09
CHONGQING ACAD OF ANIMAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, although there are reports on the detection of Salmonella based on the loop-mediated isothermal amplification technique, there is no report on the detection of Salmonella enteritidis based on the loop-mediated isothermal amplification technique.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] 1. A detection kit for Salmonella enteritidis based on loop-mediated isothermal amplification technology, consisting of the following reagents:

[0038] a. The upstream internal primer FIP aqueous solution and the downstream internal primer BIP aqueous solution with a concentration of 5 μmol / L, the upstream external primer F3 aqueous solution and the downstream external primer B3 aqueous solution with a concentration of 20 μmol / L: the primer sequences are as follows:

[0039] Upstream inner primer FIP: 5'-ttccagaacatgctcgctgc-ttgatgtggttggttcgtc-3'

[0040] Downstream internal primer BIP: 5'-tatagctcattctgacctctaagcc-gatatactccctgaatctgagaa-3';

[0041] Upstream outer primer F3: 5'-tagccaaaagaaaaccgcc-3';

[0042] Downstream outer primer B3: 5'-caggctgattactaaacctt-3';

[0043] b. An aqueous solution of Bst DNA polymerase with a concentration of 8 U / μl;

[0044] c, 10× thermal polymerization reaction buffer: Tris-HCl with a concentration of 250mmol / L and a pH of 8.8,...

Embodiment 2

[0055] 1. The Salmonella Enteritidis detection kit based on the loop-mediated isothermal amplification technology 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:

[0056] Upstream inner primer FIP: 5'-ctgagaaagaaaaactcattgaccg-ctggaaagcctctttatatagct-3';

[0057] Downstream internal primer BIP: 5'-aaggtttagtaaatcagcctgttgt-atgacaacttcgaaggtgg-3';

[0058] Upstream outer primer F3: 5'-tagccaaaagaaaaccgcc-3';

[0059] Downstream outer primer B3: 5'-tcgttcttctggtacttacg-3'.

[0060] 2. The method for detecting Salmonella enteritidis using the Salmonella enteritidis detection kit based on the loop-mediated isothermal amplification technology is the same as the method described in Example 1. It was found that the colors of the control groups 1 to 8 and the blank control group were both Yellow means that it does not contain Salmonella Enteritidis; the color of the experimental gr...

Embodiment 3

[0062] 1. The Salmonella Enteritidis detection kit based on the loop-mediated isothermal amplification technology 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:

[0063] Upstream inner primer FIP: 5'-catgctcgctgcacaaaagc-gagaggcggtttgatgtg-3';

[0064] Downstream inner primer BIP: 5'-ctggaaagcctctttatatagctca-tgatatactccctgaatctgaga-3';

[0065] Upstream outer primer F3: 5'-gggaggagctttagccaa-3';

[0066] Downstream outer primer B3: 5'-atggtgagcagacaacag-3'.

[0067] 2. The method for detecting Salmonella enteritidis using the Salmonella enteritidis detection kit based on the loop-mediated isothermal amplification technology is the same as the method described in Example 1. It was found that the colors of the control groups 1 to 8 and the blank control group were both Yellow means that it does not contain Salmonella Enteritidis; the color of the experimental group turns gr...

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Abstract

The invention discloses salmonella enteritidis detection reagent kit and method based on a loop-mediated isothermal amplification technology, wherein two specific inner primers and two specific outer primers are designed by the reagent kit according to the six specific regions of a salmonella enteritidis Sdf I gene conserved region, thereby ensuring the specificity of loop-mediated isothermal amplification and the reliability of a detecting result; in the invention, salmonella enteritidis is detected on the basis of the loop-mediated isothermal amplification technology, a target sequence can be rapidly, efficiently and specifically amplified under an isothermal condition, the operation is simple and convenient, expensive instruments and reagents are not needed, an amplified product is directly developed through fluorescent dye, a result can be judged by naked eyes, and the detection cost is low; and the reagent kit and the method can specifically distinguish serum specific type salmonella enteritidis and other serum type salmonella and are particularly suitable for detection and application in medium and small units and fields.

Description

technical field [0001] The invention relates to a biological detection kit and method, in particular to a detection kit and method for Salmonella enteritidis based on loop-mediated isothermal amplification technology. Background technique [0002] Salmonella (Salmonella) is a large group of Gram-negative bacilli with similar morphology, biochemical properties and antigenic structure. At present, there are more than 2,000 serotypes of Salmonella, and 161 serotypes have been found in my country, but only 40 to 50 serotypes are often isolated from humans and animals, mainly Salmonella typhi (Salmonella Typhi), A , Salmonella Paratyphi B, C, Salmonella Typhimurium, Salmonella Choleraesuis, Salmonella Enteritidis, Salmonella Anatum duck, Salmonella Gallinarum chicken Salmonella pullorum (Salmonella Pullorum), Salmonella Dublin (Salmonella Dublin), etc., among which Salmonella typhi, Salmonella choleraesuis and Salmonella enteritidis are the most common. [0003] Salmonella enteri...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12R1/42
Inventor 杨金龙杨松全程安春付利芝杨睿杨柳沈克飞
Owner CHONGQING ACAD OF ANIMAL SCI
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