PCR detection method of Salmonella typhimurium, nucleic acid and primer pair thereof

A Salmonella typhimurium detection method technology, applied in the field of Salmonella typhimurium PCR detection method and nucleic acid and primer pairs therein, can solve the problem that the PCR detection method of Salmonella typhimurium serotype PCR detection method has not been invented, the nucleic acid and primer, detection target point specificity is not strong, Increase detection complexity and other issues to achieve the effect of practicability, simple result judgment, and short detection time

Active Publication Date: 2012-07-18
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But there are only few PCR methods for Salmonella typhimurium serotype detection, and the specificity of the detection target is not strong, such as Lim YH, Hirose K, Izumiya H, etc. in "Japanese Journal of Infectious Diseases" (Japanese Journal of Infectious Diseases) 2003 Volume 56, Issue 4, pages 151-155, published in the article entitled "Multiplex polymerase chain reaction assay for selective detection of Salmonella enterica serovar typhimurium" (selective detection of Salmonella typhimurium by multiplex PCR method), according to typhoid fever The serotype O4:Hi:1,2 of Salmonella adopts the corresponding flagellar gene encoding the H antigen? iC and ? jB and O antigen gene rfbJ form multiplex PCR to detect Salmonella typhimurium, which increases the complexity of detection, the interference between primers easily affects the improvement of detection sensitivity, and the strains with similar serotypes will bring difficulties to the judgment of the final result
[0003] Find through literature search to prior art, as yet invention and the Salmonella typhimurium serotype PCR detection method of the present invention, nucleic acid and the relevant report of primer

Method used

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  • PCR detection method of Salmonella typhimurium, nucleic acid and primer pair thereof
  • PCR detection method of Salmonella typhimurium, nucleic acid and primer pair thereof
  • PCR detection method of Salmonella typhimurium, nucleic acid and primer pair thereof

Examples

Experimental program
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Embodiment 1

[0021] Establishment of PCR detection method for serotypes of Salmonella typhimurium

[0022] Step 1, designing primers according to the conserved sequence in the genomic DNA sequence of Salmonella typhimurium

[0023] Find the specific STM4495 gene from the Salmonella typhimurium genome DNA sequence, and use it as the detection target gene of Salmonella typhimurium, the gene sequence is shown in SEQ ID NO: 1;

[0024] Input the DNA sequence of the restriction endonuclease gene into the primer design software Primer Premier 5.0 to design primers, set the GC% range to 40-60%, and the product size range to 100-1000bp, and select primers from the alternative primer pairs , the primer sequences are as follows (primers were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd.):

[0025] STM4-L: 5'-GGTGGCAAGGGAATGAA-3' (SEQ ID NO: 2);

[0026] STM4-R: 5'-CGCAGCGTAAAGCAACT-3' (SEQ ID NO: 3);

[0027] Step 2, DNA template preparation

[0028] Inoculate Salmon...

Embodiment 2

[0041] Detection of suspected strains of Salmonella typhimurium

[0042] Using the PCR detection method for Salmonella typhimurium established in Example 1, 61 suspected strains of Salmonella isolated from food samples were detected. The food samples were collected in suburban supermarkets and bazaars in Shanghai. For sample processing and isolation of suspected strains, refer to the national standard GB / T 4789.4-2008.

[0043] From it, 9 suspected bacterial strains were detected as positive results, and these 9 suspected bacterial strains were identified as O4:Hi:5 by the Salmonella diagnostic serum (Lanzhou Institute of Biological Products), and it was determined that it was Salmonella typhimurium (see the product manual for the serum identification steps and national standard GB / T 4789.4-2008), other suspected strains were identified as not Salmonella typhimurium. This example proves that the PCR detection method for Salmonella typhimurium has very high reliability.

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Abstract

The invention relates to a PCR detection method of Salmonella typhimurium, nucleic acid and a primer pair thereof, belonging to the technical field of the safety inspection of foods; the detection method comprises the following steps: designing an amplification primer according to a conserved sequence SEQ ID NO:1 in the genome DNA sequence of the Salmonella typhimurium; extracting a sample DNA and amplifying by a PCR method; and detecting an amplified product by gel electrophoresis and judging whether the sample contains the Salmonella typhimurium or not; the judgment comprises the following concrete steps: if a corresponding single-amplification strip occurs in an electrophoresis result, showing that the sample contains the Salmonella typhimurium; if the corresponding amplification strip does not occur, showing that the sample does not contain the Salmonella typhimurium; the invention also relates to the nucleic acid of which the base sequence is shown as SEQ ID NO:1; and the invention also relates to the primer pair, in particular to the nucleic acid of which the base sequence is shown as SEQ ID NO:2 and SEQ ID NO:3. By adopting the detection method to detect the Salmonella typhimurium, the detection time is short, the cost is low, the detection result is specific, and the result judgment is simple.

Description

technical field [0001] The invention relates to a PCR detection method in the technical field of food safety inspection and a nucleic acid and a primer pair therein, in particular to a PCR detection method for Salmonella typhimurium and a nucleic acid and a primer pair therein. Background technique [0002] Salmonella typhimurium (Salmonella Typhimurium) was isolated from mice by Loffler in 1892 and is one of the most common serotypes of Salmonella infection. Salmonella typhimurium belongs to group B of the genus Salmonella, and its biological characteristics are similar to those of Salmonella typhi. It is a Gram-negative bacterium that does not form spores, has no capsule, but has flagella. The fungus formed three varieties in the evolution process, namely the Copenhagen variety, the Binns variety and the O-type variety. Salmonella typhimurium has strong resistance to the external environment, can reproduce rapidly at room temperature, and can survive in low temperature an...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12R1/42
Inventor 史贤明刘斌何晓华施春雷陈婧
Owner SHANGHAI JIAO TONG UNIV
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