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Method for synthesizing quenching fluorescence RNA marker and method for detecting food-borne pathogenic bacteria

A technology of food-borne pathogenic bacteria and fluorescent markers, which is applied in the field of quenching fluorescent RNA marker synthesis, and can solve the problems of unreported nucleic acid of food-borne pathogenic bacteria

Active Publication Date: 2019-09-17
JIANGSU UNIV
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Problems solved by technology

[0003] At present, the rapid detection method of nucleic acid of food-borne pathogenic bacteria using up-conversion fluorescent nanotechnology has not been reported yet.

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  • Method for synthesizing quenching fluorescence RNA marker and method for detecting food-borne pathogenic bacteria
  • Method for synthesizing quenching fluorescence RNA marker and method for detecting food-borne pathogenic bacteria

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Embodiment Construction

[0017] In order to make the objectives, technical solutions and advantages of the present invention clearer, the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention, but not to limit the present invention.

[0018] The present invention is applicable to the detection of food-borne pathogenic bacteria. In the present embodiment, only Escherichia coli (E.coli) is used as an example, and the specific operation steps are as follows:

[0019] Upconversion nanoparticles with amino modifications were prepared by a solvothermal method. The specific process is as follows: the NH 4 F (6.24 mmol) was dissolved in 12 mL of EG. Take NaCl (1mmol), PEI (0.5g), Gd (NO 3 ) 3 (0.8mmol), Yb(NO 3 ) 3 (0.17mmol) and Er(NO 3 ) 3 (0.03 mmol) was dissolved in EG (38 mL) and stirred magnetically for 30 min. W...

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Abstract

The invention discloses a method for synthesizing a quenching fluorescence RNA marker and a method for detecting food-borne pathogenic bacteria. The synthesis method comprises the following steps: synthesizing up-conversion nanoparticles with amino modification by adopting a one-step solvothermal method; combining specific RNA with a quencher to synthesize an RNA fluorescent marker; and connecting the RNA fluorescent marker with the up-conversion nanoparticles with amino modification by adopting a glutaraldehyde crosslinking method so as to obtain the quenching fluorescence RNA marker. In addition, the quenching fluorescence RNA marker is applied to the detection of food-borne pathogenic bacteria. The detection comprises the following steps: accurately cutting a specific nucleic acid target in trace pathogenic bacteria cells by using nuclease Cas13a, shearing the quenching fluorescence RNA marker by utilizing the incidental cutting effect of the nuclease, and releasing fluorescence which can be detected; and collecting fluorescence spectrum data, and constructing a quantitative detection model of fluorescence intensity change values and different amounts of food-borne pathogenic bacteria nucleic acid targets so as to realize nano fluorescence trace detection of food-borne pathogenic bacteria nucleic acid.

Description

technical field [0001] The invention belongs to the technical field of foodborne pathogenic bacteria detection, in particular to a method for synthesizing a quenching fluorescent RNA marker and a method applied to the detection of foodborne pathogenic bacteria. Background technique [0002] Food-borne pathogens are one of the main causes of food-borne diseases, and are important to the world's food safety, which has seriously threatened human health. The detection of food-borne pathogens is an important means of food safety assurance. With the rapid development of nucleic acid detection technology, rapid detection methods for various food-borne pathogens have emerged one after another. Commonly used are polymerase chain reaction and its derivative technology, nucleic acid isothermal amplification technology, oligonucleotide microarray technology and immunomagnetic cell separation technology. Although these existing methods have their own advantages, they have shortcomings ...

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Application Information

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IPC IPC(8): C12Q1/6823C12Q1/6806
CPCC12Q1/6823C12Q1/6806C12Q2521/301C12Q2563/107C12Q2563/155Y02A50/30
Inventor 陈全胜李欢欢刘蕊吕鹏欧阳琴焦天慧朱家骥郭志明
Owner JIANGSU UNIV
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