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A method for the detection of Pseudomonas aeruginosa based on aptamer fluorescence sensing

A Pseudomonas aeruginosa, fluorescence sensing technology, applied in fluorescence/phosphorescence, measuring device, material analysis by optical means, etc., can solve the problems of high cost, complex process, time-consuming, etc., and achieve the effect of easy production

Inactive Publication Date: 2020-10-09
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods still have problems such as high cost of preparing materials, complicated process, and time-consuming.

Method used

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  • A method for the detection of Pseudomonas aeruginosa based on aptamer fluorescence sensing
  • A method for the detection of Pseudomonas aeruginosa based on aptamer fluorescence sensing
  • A method for the detection of Pseudomonas aeruginosa based on aptamer fluorescence sensing

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1 Real-time fluorescence detection of Pseudomonas aeruginosa live bacteria

[0029] A detection method for Pseudomonas aeruginosa, characterized in that it comprises the steps:

[0030] (1) Bacterial culture: resuscitate the purchased Pseudomonas aeruginosa freeze-dried powder, streak and inoculate them on the Lb nutrient agar plate and Lb liquid medium respectively, place the Lb nutrient agar plate in a refrigerator at 20°C for cultivation, and liquid culture The base was placed in a constant temperature incubator, set the temperature at 48°C, and the rotation speed was 150r / min, and cultivated for 48h. After shaking for a certain period of time, the culture is harvested and then used for detection. The obtained live bacterial suspension can be used directly or stored at 4°C;

[0031] (2) Count Pseudomonas aeruginosa colonies using the coating method, and calculate the bacterial concentration (CFU / mL) used for detection;

[0032] (3) The hairpin structure ...

Embodiment 2

[0037] Example 2 Real-time fluorescence detection of antibiotic inactivation of Pseudomonas aeruginosa

[0038] A detection method for antibiotic inactivation of Pseudomonas aeruginosa, characterized in that it comprises the steps:

[0039] (1) Bacterial culture: resuscitate the freeze-dried powder of Pseudomonas aeruginosa, streak and inoculate them on Lb nutrient agar plate and Lb liquid medium respectively, place the Lb nutrient agar plate in a refrigerator at 20°C for cultivation, and place the liquid medium in The constant temperature incubator is adjusted, the temperature is set at 48°C, the rotation speed is 150r / min, and the incubation is carried out for 48h. After shaking for a certain period of time, the cultures were harvested and used for assays. The resulting live bacterial suspension can be used directly or stored at 4°C;

[0040] (2) Count Pseudomonas aeruginosa colonies using the coating method, and the calculated colony concentration is 1.11*10 9 (CFU / mL); ...

Embodiment 3

[0047] Real-time fluorescence detection of Pseudomonas aeruginosa when embodiment 3 has interfering bacteria to exist

[0048] The detection method of Pseudomonas aeruginosa when interfering bacteria exists, is characterized in that comprising the steps:

[0049] (1) Bacterial culture: resuscitate the lyophilized powder of Pseudomonas aeruginosa, streak and inoculate them on Lb nutrient agar plate and Lb liquid medium respectively, place the Lb nutrient agar plate in a refrigerator at 20°C for cultivation, and inoculate Escherichia coli (BNCC336685 ) was recovered from the cryopreserved liquid, inoculated in Lb liquid medium, and the liquid medium was placed in a constant temperature incubator with a temperature of 48°C and a rotation speed of 150r / min, and cultured for 48 hours. After shaking for a certain period of time, the cultures were harvested and used for assays. The resulting live bacterial suspension can be used directly or stored at 4°C;

[0050] (2) Count the col...

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Abstract

The invention discloses a pseudomonas aeruginosa detection method based on aptamer fluorescence sensing. An aptamer is combined with a fluorescence signal strategy, a simple fluorescence biosensor (aptamer fluorescence sensor) with strong selectivity is developed and used for detecting the pseudomonas aeruginosa. The aptamer and the fluorescence signal strategy are combined, a specificity probe hairpin structure DNA modified BHQ-2 and cy5 is mixed and hybridized with the single-stranded DNA aptamer; after adding thallus in the mixed solution, the bacteria can be combined with the single-stranded DNA aptamer specificity, the single-stranded DNA and the hairpin structure DNA are separated, the hairpin structure is recovered, and the fluorescence of the cy5 is further quenched. The DNA aptamer, the viable bacteria and the fluorescence signal are skillfully combined to construct an extremely simple system for detecting the viable bacteria. The detection method disclosed by the invention isnot only simple, quick, sensitive and low in equipment requirement, but also can be applied to the online monitoring of the viable bacteria.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to a method for detecting Pseudomonas aeruginosa based on aptamer fluorescence sensing. Background technique [0002] In recent years, bacterial infections have been one of the problems that plague our public health security, and one of the most important global risks is some bacterial infections with high potential. Nonetheless, multiple challenges and obstacles remain in the prevention and control of bacterial infections. The increasing population and complexity of bacterial infections are leading to synergistic effects. Among them, Pseudomonas aeruginosa is a nonfermenting Gram-negative bacillus that can cause fatal infections in a variety of immunocompromised diseases, such as cancer, cystic fibrosis (CF) and burns. Recently, the World Health Organization (WHO) declared Pseudomonas aeruginosa one of the 12 deadliest superbugs and its multidrug resistance must be addressed a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569G01N21/64
CPCG01N21/6486G01N33/56911
Inventor 陈宪徐礼明
Owner FUZHOU UNIV
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