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A target gene, specific primer pair, detection method and kit for detecting Salmonella delphi

A Salmonella and primer pair technology, applied in the field of molecular biology and biological detection, can solve the problems of long detection period and complex determination of Salmonella Delpy detection results, and achieve the effects of short detection period, reliable detection results and simple detection process.

Active Publication Date: 2020-11-17
ANHUI SCI & TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the deficiencies in the prior art, the present invention provides a target gene, a specific primer pair, a detection method and a kit for detecting Salmonella Delphi, which solves the problems of complex determination of Salmonella Delphi detection results and long detection period in the prior art. technical problem

Method used

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  • A target gene, specific primer pair, detection method and kit for detecting Salmonella delphi
  • A target gene, specific primer pair, detection method and kit for detecting Salmonella delphi
  • A target gene, specific primer pair, detection method and kit for detecting Salmonella delphi

Examples

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Effect test

Embodiment 1

[0034] Specificity evaluation of different bacterial species by PCR detection method

[0035] 1. Screening of serotype-specific genes of Salmonella delbei

[0036]In the NCBI database, the whole genome sequence of Salmonella delphi CVM40386 strain (NZ_JYYM01000031) was obtained. According to the method of comparing genomes, each gene of the genome of the bacteria was compared using the Blast program of the NCBA website, and the homology with the serotype was selected. Higher (E value = 0, Qμery cover = 0%) and low homology (Qμery cover < 10%) genes with other microorganisms as the quasi-specific genes of Salmonella delbei. A total of 12 quasi-specific genes of Salmonella delbes were obtained by this method, and then multiple pairs of primers were designed for each gene, and the specificity was verified by using other serotypes of Salmonella preserved in the laboratory. According to the PCR results, the RU61_00441, RU61_00445, RU61_00447, RU61_RS09205, and RU61_RS06985 genes w...

Embodiment 2

[0048] Sensitivity Evaluation of PCR Detection Method to Different Template Concentrations

[0049] The screening and primer design steps of Salmonella delbes serotype-specific genes are as in Example 1.

[0050] Preparation of DNA template: Pick a single colony of Salmonella delbee and transfer it to 30ml of LB liquid medium, culture overnight at 37°C. The bacterial genomic DNA mini-extraction kit produced by Shanghai Sangon Bioengineering Technology Service Co., Ltd. was used to extract the total DNA of Salmonella delphi, and the concentration was determined to be 265.8ng / μl. Make a 10-fold gradient dilution with sterile water, and dilute 7 gradients in total as PCR templates.

[0051] Establishment of PCR amplification reaction system: establishment of PCR detection system: 25 μl reaction system includes: 12.5 μl of 2×Master, 1 μl of 10 μM primer pair, 5 μl of template, supplemented with ddH2O to 25 μl. The reaction program of PCR was designed: pre-denaturation at 94°C fo...

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Abstract

The invention provides a target gene, a specific primer pair, a detection method and a kit for detecting salmonella derby. The specific primer is designed according to five salmonella derby specific detection targets, and the detection targets are high in stability and high in specificity. The detection method comprises the steps of extracting sample DNA, and carrying out PCR amplification; detecting amplification products through gel electrophoresis; and comparing strips after the electrophoresis. Through experimental comparative analysis, the method has the characteristics of being high in oneness, reliable in detection result and simple in result judgment, and can be widely applied to the field of food sanitation.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and biological detection, and relates to a target gene, a pair of specific primers, a detection method and a kit for detecting Salmonella delphi. Background technique [0002] Salmonella Derby (Salmonella Derby) is one of the main Salmonella serotypes that cause human infection, and the main host of this serotype strain is humans. Salmonella delphi can survive in livestock and poultry. Once the feces of livestock and poultry or raw meat carrying the pathogen contaminates food and processing sites, the bacterium will grow and reproduce rapidly in an environment with a suitable temperature. When the number of bacteria in the food Reaching a certain amount, food poisoning will be caused after being eaten, endangering people's physical and mental health, and even life-threatening in severe cases. Zheng Huaying, Zhou Dunjin, etc. in 2003 "A Survey on the Outbreak of Intestinal Infectious Di...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/10C12N15/11C12R1/42
CPCC12Q1/689
Inventor 翟立公王俊颖王蓓蓓王丽郭元新杨剑婷杜传来
Owner ANHUI SCI & TECH UNIV