Method for detecting mycobacterium tuberculosis by in-situ fluorescent loop-mediated isothermal nucleic acid amplification technology and kit

Abstract

The invention discloses a method for detecting mycobacterium tuberculosis by in-situ fluorescent loop-mediated isothermal nucleic acid amplification technology and a kit. Four specific primers are designed in a mycobacterium tuberculosis gyrB gene conservation area; and the mycobacterium tuberculosis can be specifically detected by using the four primers and combining the in-situ fluorescent loop-mediated isothermal nucleic acid amplification technology at the same time. The method realizes in-situ detection of a sample, does not need to increase bacteria, has high specificity and can finish amplification in less than 2 hours; the lowest detection limit reaches 10CFU / ml; and the detection rate of the sample reaches 99 percent. The method and the kit for identifying the mycobacterium tuberculosis are simple and convenient.

Description

technical field [0001] The invention relates to a microorganism detection reagent and a method, in particular to a method and a kit for detecting mycobacterium tuberculosis by an in-situ fluorescent ring-mediated constant temperature nucleic acid amplification technique. Background technique [0002] At present, tuberculosis is the number one killer of infectious diseases in the world today, and it is the infectious disease with the highest single-cause fatality rate. Its main pathogenic bacteria is Mycobacterium tuberculosis. According to the statistics of the World Health Organization, there are 20 million tuberculosis patients in the world, about 9 million new cases occur every year, and 3 million people die every year. About 1 / 3 of the population has latent Mycobacterium tuberculosis infection. Tuberculosis is on the rise due to population growth, immigration, tourism, drug resistance, the prevalence of human immunodeficiency virus infection, drug abuse, alcohol abuse, a...

AI Technical Summary

Problems solved by technology

Although the disclosed LAMP technology has high sensitivity and can efficiently amplify nucleic acid under isothermal conditions, the existing technology also has deficiencies, mainly: (1) the requirements for special primers are extremely harsh, which limits the wide application of this method; (2) Cells or tissues need to be crushed to extract DNA or RNA, and viruses or genes in cells or tissues cannot be directly located; (3) In order to meet the amount of DNA required for amplification, the number of pathogenic bacteria must be multiplied; ( 4) Judging the result only by the milky white precipitate generated by the reaction is not clear and accurate enough, and it is not easy to realize the miniaturization of the reaction, which limits its further development

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