Method for detecting salmonella enteritidis in food

A technology of Salmonella Enteritidis and a detection method, applied in the field of bacterial detection, can solve the problems of time-consuming, unfavorable promotion and use of basic units, long detection time, etc., and achieves the effects of simple pretreatment process, low sample detection cost, and long storage time.

Inactive Publication Date: 2017-05-31
WUXI X RES PROD DESIGN & RES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although this method is accurate, it is laborious, costly, time-consuming, and cannot detect products with a short shelf life
At present, the method for detecting Salmonella takes a long time and costs a lot, which is not conducive to the promotion and use in grassroots units

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] A method for detecting Salmonella Enteritidis in food, adopting a constant temperature amplification method, taking the specific sequence of the Salmonella Enteritidis clpP gene as a target sequence, SYBER Green I as a fluorescent dye, and then performing fluorescence quantitative detection and analysis; the specific process includes the following steps:

[0036] (1) Preprocessing:

[0037] In a sterile environment, the food was added to BPW buffered peptone water, the culture temperature was 32°C, and the culture time was 22 hours. After the culture was over, it was washed with normal saline repeatedly, and then genomic DNA and RNA were extracted;

[0038] (2) Construct a constant temperature amplification reaction system:

[0039] DNA template 2 μL, SYBER Green I fluorescent dye 0.5 μL, 10× Buffer 3.0 μL, DNA polymerase 7U, ammonium chloride solution 0.5 μL, aluminum chloride solution 0.5 μL, dNTP 3 μL, DNA primer 3 μL, carbonic anhydrase 2U, deoxygenation Inosine so...

Embodiment 2

[0053] A method for detecting Salmonella Enteritidis in food, adopting a constant temperature amplification method, taking the specific sequence of the Salmonella Enteritidis clpP gene as a target sequence, SYBER Green I as a fluorescent dye, and then performing fluorescence quantitative detection and analysis; the specific process includes the following steps:

[0054] (1) Preprocessing:

[0055]In a sterile environment, the food was added to BPW buffered peptone water, the culture temperature was 32°C, and the culture time was 22 hours. After the culture was over, it was washed with normal saline repeatedly, and then genomic DNA and RNA were extracted;

[0056] (2) Construct a constant temperature amplification reaction system:

[0057] DNA template 2 μL, SYBER Green I fluorescent dye 0.5 μL, 10× Buffer 3.0 μL, DNA polymerase 7U, ammonium chloride solution 0.5 μL, aluminum chloride solution 0.5 μL, dNTP 3 μL, DNA primer 3 μL, carbonic anhydrase 2U, deoxygenation Inosine sol...

Embodiment 3

[0071] A method for detecting Salmonella Enteritidis in food, adopting a constant temperature amplification method, taking the specific sequence of the Salmonella Enteritidis clpP gene as a target sequence, SYBER Green I as a fluorescent dye, and then performing fluorescence quantitative detection and analysis; the specific process includes the following steps:

[0072] (1) Preprocessing:

[0073] In a sterile environment, the food was added to BPW buffered peptone water, the culture temperature was 33°C, and the culture time was 23 hours. After the culture was over, it was washed repeatedly with normal saline, and then genomic DNA and RNA were extracted;

[0074] (2) Construct a constant temperature amplification reaction system:

[0075] DNA template 2 μL, SYBER Green I fluorescent dye 0.5 μL, 10× Buffer 3.0 μL, DNA polymerase 7U, ammonium chloride solution 0.5 μL, aluminum chloride solution 0.5 μL, dNTP 3 μL, DNA primer 3 μL, carbonic anhydrase 2U, deoxygenation Inosine so...

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PUM

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Abstract

The invention discloses a method for detecting salmonella enteritidis in food. An isothermal amplification method is adopted, a clpP gene specific sequence of the salmonella enteritidis is taken as a target sequence, and SYBER Green I is used as a fluorescent dye; then, fluorescence quantitative detection analysis is carried out; the concrete process comprises the following steps: (1) carrying out pretreatment; (2) establishing an isothermal amplification reaction system; (3) carrying out a fluorescent quantitation reaction; (4) carrying out detection analysis. A primer designed by the method is good in detection specificity and high in sensitivity, and is very suitable for detecting the salmonella enteritidis in the food.

Description

technical field [0001] The invention relates to a method for detecting bacteria, in particular to a method for detecting Salmonella enteritidis in food. Background technique [0002] Salmonella belongs to the Enterobacteriaceae Salmonella genus. According to its antigen structure and biochemical tests, there are currently more than 2,000 serotypes, among which Salmonella typhimurium, Salmonella enteritidis and Salmonella choleraesuis are more common. The bacterium is a gram-negative bacillus, aerobic, does not produce spores, has no capsule, and most of them have flagella and can move. It has strong resistance to the outside world. It can survive for several months in water and soil, 1 to 2 months in feces, and survive the winter in frozen soil. It is not heat resistant, it will die at 55°C for 1 hour or 60°C for 10 to 20 minutes, and it can be killed within 5 minutes by 5% carbolic acid or 1:500 liters of mercury. Such bacteria can be found in the intestinal cavity and vi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/10C12R1/42
CPCC12Q1/689C12Q1/6844C12Q2563/107C12Q2545/114
Inventor 吴敏芳徐静赵春城胡勇蒋韦艳刘金杰
Owner WUXI X RES PROD DESIGN & RES
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