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Multiple PCR detection primers of escherichia coli, pseudomonas aeruginosa and staphylococcus aureus, kit and detection method

A Pseudomonas aeruginosa detection kit technology, applied in the field of microbial detection, achieves the effects of strong detection specificity, saving detection cost and time, and improving efficiency and accuracy

Inactive Publication Date: 2017-02-15
GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY
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  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Multiplex PCR is a new type of DNA amplification technology improved and developed on the basis of conventional PCR. It is to add multiple pairs of primers to amplify DNA fragments of multiple entries in the same reaction system at the same time. Using this technology, a variety of pathogenic microorganisms can be amplified. This technology has been used in food and drinking water, but it is rare in the field of cosmetics. Therefore, it is urgent to establish a rapid, efficient and simultaneous detection of multiple pathogenic microorganisms. Microbiological detection method for cosmetics

Method used

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  • Multiple PCR detection primers of escherichia coli, pseudomonas aeruginosa and staphylococcus aureus, kit and detection method
  • Multiple PCR detection primers of escherichia coli, pseudomonas aeruginosa and staphylococcus aureus, kit and detection method
  • Multiple PCR detection primers of escherichia coli, pseudomonas aeruginosa and staphylococcus aureus, kit and detection method

Examples

Experimental program
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Effect test

Embodiment 1

[0026] Example 1: Design and Screening of Multiplex PCR Detection Primers

[0027] 1. Primer design

[0028] The Escherichia coli alkaline phosphatase gene (phoA gene), Pseudomonas aeruginosa outer membrane protein gene (oprL gene) and Staphylococcus aureus thermostable nuclease gene (nuc gene) were used as specific detection target sequences respectively, and primers were carried out Design and screen out 3 pairs of primers with high specificity as follows:

[0029] For Escherichia coli phoA gene: phoA-F: 5'-GTTTCTACCGCAGAGTTG-3' (as shown in SEQ ID NO.1), phoA-R: 5'-GACTATGACCAGCGTGTT-3' (as shown in SEQ ID NO.2), The length of the amplified fragment is 663bp.

[0030] For Pseudomonas aeruginosa oprL gene: oprL-F: 5'-GGCGTGCTGATGCTCGTA-3' (as shown in SEQ ID NO.3), oprL-R: 5'-CGCTGACCGCTGCCTTTC-3' (as shown in SEQ ID NO.4 ), the length of the amplified fragment was 444bp.

[0031] For Staphylococcus aureus nuc gene: nuc-F: 5'-ACATAAAGAACCTGCGAC-3' (as shown in SEQ ID NO....

Embodiment 2

[0044] Example 2: Detection of 3 kinds of pathogenic bacteria in cosmetics

[0045] (1) Preparation of artificially polluted samples: Seven different types of cosmetic samples submitted by a company for inspection are moisturizing milk, lotion, face cream, facial cleanser, toner, liquid foundation, and essence, according to the "Hygienic Standards for Cosmetics" ( 2007 version) in the microbial detection method, no microbial contamination was found, and artificial random pathogenic bacteria contamination was carried out. Escherichia coli ATCC8039 and Pseudomonas aeruginosa ATCC9027 were added to the moisturizing milk, and Pseudomonas aeruginosa ATCC9027 and grape aureus were added to the lotion Coccus ATCC6538, Escherichia coli ATCC8039 and Staphylococcus aureus ATCC6538 were added to the cream, Escherichia coli ATCC8039, Pseudomonas aeruginosa ATCC9027 and Staphylococcus aureus ATCC6538 were added to the cleanser, Pseudomonas putida and Staphylococcus saprophyticus were added ...

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Abstract

The invention discloses multiple PCR detection primers of escherichia coli, pseudomonas aeruginosa and staphylococcus aureus, a kit and a detection method. Specific primers are designed by means of combined utilization of escherichia coli alkaline phosphatase gene, pseudomonas aeruginosa outer membrance proteins gene and staphylococcus aureus heat-resistant nuclease gene sequences, multiple PCR detection is conducted on a sample by means of the specific primers according to the method, and specific detection of the escherichia coli, the pseudomonas aeruginosa and the staphylococcus aureus in the sample can be achieved at a time; the detection time is short, operation is easy and convenient, the detection efficiency can be effectively improved, the requirement of rapid detection is met, the sensitivity is high, the specificity is high, and the multiple PCR detection primers of the escherichia coli, the pseudomonas aeruginosa and the staphylococcus aureus, the kit and the detection method can be applied to microbiological detection work of cosmetics.

Description

[0001] Technical field: [0002] The invention belongs to the field of microorganism detection, and in particular relates to multiple PCR detection primers, a kit and a detection method for Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus. [0003] Background technique: [0004] Cosmetics contain nutrient-rich protein, fat, vitamins, inorganic salts, etc., which are extremely conducive to the growth and reproduction of microorganisms. The growth and reproduction of microorganisms can cause spoilage of cosmetics, and can also produce toxins or metabolites. As allergens or irritants, these foreign bodies may cause sensitization or irritation to the application site, causing various types of cosmetic skin diseases, such as contact dermatitis, acne, hair damage, photosensitive dermatitis, and abnormal skin pigmentation. Therefore, " Hygienic Standards for Cosmetics (2007 Edition) stipulates that fecal coliforms, Pseudomonas aeruginosa and Staphylococcus aureus sh...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/14C12Q1/10C12Q1/04C12N15/11C12R1/19C12R1/385C12R1/445
CPCC12Q1/689C12Q1/686C12Q2600/16C12Q2537/143
Inventor 文霞杨秀茳谢小保孙廷丽周少璐
Owner GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY
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