CRISPR/Cas13a based food-borne pathogenic bacterium nucleic acid nano fluorescence trace detection method

Abstract

The invention discloses a CRISPR / Cas13a based food-borne pathogenic bacterium nucleic acid nano fluorescence trace detection method. The method includes taking food-borne pathogenic bacterium nucleicacid as a study object, and preparing food-borne pathogenic bacterium nucleic acid targets, food-borne pathogenic bacterium crRNA, and nuclease Cas13a protein purification and quenching fluorescent RNA reporting markers; utilizing nuclease Cas13a to perform accurate cutting on specific nucleic acid targets in trace pathogenic bacterium cells, and cutting the quenching fluorescent RNA reporting markers by utilizing the incidental cutting effects of the nuclease to release fluorescence that can be detected; and collecting fluorescence spectrum data with the help of a fluorescence spectrum system, obtaining the fluorescence intensity value of the maximum absorption peak of an up-conversion fluorescent nanomaterial, and constructing a quantitative determination model of up-conversion nano fluorescence intensity and food-borne pathogenic bacterium nucleic acid target content so that the rapid detection on food-borne pathogenic bacterium nucleic acid nano fluorescence trace can be realized.The method is short in detection period, strong in specificity and high in sensitivity.

Description

technical field [0001] The invention belongs to the technical field of trace detection of food-borne pathogenic bacteria, and in particular relates to a CRISPR / Cas13a-based nanofluorescent trace detection method of nucleic acid of food-borne pathogenic bacteria. Background technique [0002] Food-borne pathogenic bacteria are one of the main causes of food-borne diseases, are important to the world's food safety, and have seriously threatened human health. The detection of foodborne pathogenic bacteria is an important means of food safety assurance. With the rapid development of nucleic acid detection technology, rapid detection methods for various foodborne pathogens have emerged one after another. Commonly used are polymerase chain reaction and its derivative technology, nucleic acid constant temperature amplification technology, oligonucleotide microarray technology and immunomagnetic cell separation technology. Although these existing methods have their own advantages,...

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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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