Salmonella enteritidis nucleic acid standard sample as well as building method and application thereof

A technology of Salmonella Enteritidis and standard sample, which is applied in the preparation of Salmonella Enteritidis nucleic acid standard sample and the application field in detecting Salmonella Enteritidis, and achieves the effect of broad market, great economic benefits and social benefits

Inactive Publication Date: 2012-10-10
曹际娟 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem that the nucleic acid standard samples of pathogenic bacteria in food are currently in a blank state at home and abroad, the present invention mainly aims at the qualitative and quantitative detection items of the nucleic acid of the pathogenic bacteria Salmonella enteritidis in food, and researches and prepares the standard sample of nucleic acid of Salmonella enteritidis in food.

Method used

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  • Salmonella enteritidis nucleic acid standard sample as well as building method and application thereof
  • Salmonella enteritidis nucleic acid standard sample as well as building method and application thereof
  • Salmonella enteritidis nucleic acid standard sample as well as building method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Source of strains: The standard strains used in the preparation of Salmonella Enteritidis nucleic acid standard samples were purchased from the American Type Culture Collection, and the standard strain number of Salmonella Enteritidis is ATCC: 13076. figure 1 Process flow chart for the preparation of Salmonella Enteritidis (Salmonella Enteritidis) nucleic acid standard samples.

[0052] In this article, the experimental procedures, experimental reagent preparation and other methods related to genetic engineering operations, unless otherwise specified, refer to the "Molecular Cloning Experiment Guide".

[0053] Genomic nucleic acid extraction method:

[0054] According to GB / T4789.4-2008 [1] Carry out strain activation and enrichment culture. The DNA of the enrichment broth was extracted using Qiagen’s DNA extraction kit (Qiagen Blood & Cell Culture DNA Maxi Kit, Cat. No. 13362), and the genomic DNA was extracted according to its operating instructions. Prepare nuclei...

Embodiment 2

[0079] Certainty test

[0080] 1PCR analysis

[0081] a. Instrument: PE2400, American PE Company

[0082] b. Determination: Take the prepared nucleic acid standard sample and determine by PCR analysis:

[0083] Specific fragment invA gene PCR primers in Salmonella Enteritidis:

[0084] Forward primer: SEQ ID NO: 1 5'-gtgaaattat cgccacgttc gggcaa-3'

[0085] Reverse primer: SEQ ID NO: 2 5′-tcatcgcacc gtcaaagaac c-3′

[0086] Reaction conditions:

[0087] Pre-denaturation: 94°C, 3min;

[0088] Entering the cycle: denaturation at 94°C for 60s, annealing at 60°C for 60s, extension at 72°C for 60s, 35 cycles;

[0089] Termination of extension: 72°C, 7min;

[0090] The length of the amplified product was 284bp, and a specific gene fragment of Salmonella Enteritidis was amplified, which was confirmed to be Salmonella Enteritidis after sequencing.

[0091] 2 Real-time fluorescent PCR analysis

[0092] a. Instrument: ABI 7500 real-time fluorescent quantitative PCR analyzer

[...

Embodiment 3

[0166] The preparation of Salmonella Enteritidis nucleic acid standard sample can be widely used in various food detection kits as a positive standard control substance. The specific preparation method is as described in Example 1 above.

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Abstract

The invention discloses a Salmonella enteritidis nucleic acid standard sample as well as a building method and application thereof. The building method comprises the steps: strain culture, identification, preparation of a nucleic acid standard sample, drying of the nucleic acid standard sample, uniformity and stability inspection, qualitative assay, rating of the nucleic acid standard sample and other operations to obtain the Salmonella enteritidis nucleic acid standard sample. According to the invention, the condition that the pathogen nucleic acid standard sample is deficient in domestic food is solved, has important practical significance in realizing preparation technology of the pathogen nucleic acid standard sample in food and stability guarantee technology, positively developing the research and development of pathogen molecular biological detection standard sample in domestic food, and filling up the blank of the measuring field, and has a broad market in developing molecular biological detection working of pathogen in the food and food pathogen nucleic acid standard sample, and has greater economic benefits and social benefits.

Description

technical field [0001] The invention belongs to the technical field of food detection, and in particular relates to the preparation of a Salmonella enteritidis nucleic acid standard sample, its establishment method and its application in the detection of Salmonella enteritidis. Background technique [0002] Food safety is a major public health issue and an important factor restricting my country's economic development. Foodborne diseases caused by microbial contamination are the most prominent problems in my country's food safety. In recent years, with the adjustment of food industry structure, the rapid development of economic globalization and international trade, the prevention and control of foodborne diseases in my country is facing a series of new problems and challenges. [0003] The prevention and control of foodborne diseases caused by microorganisms in food has attracted the attention of countries all over the world. The detection technology of microorganisms in f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/68C12R1/42
Inventor 曹际娟于兵徐杨徐君怡
Owner 曹际娟
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