Legionella pneumophila detection kit

Abstract

The invention discloses a quick detection kit for legionella pneumophila. A pair of legionella specific primers is used for amplifying a region on a 16S rRNA (ribonucleic acid) gene; a pair of hybridization probes is designed in the amplified fragment aiming at a legionella pneumophila specific sequence; an FAM fluorophore is labeled at an anchoring probe end 3'; a BHQ1 is labeled at a quenching probe end 5'; the end 3' is phosphorylated; the legionella pneumophila in a sample is detected through real-time fluorescence PCR (polymerase chain reaction) dissolving curve analysis. The detection kit mainly comprises a DNA (deoxyribonucleic acid) extracting solution, a fluorescence PCR reaction solution, a negative contrast and a positive contrast. The detection method comprises three steps of DNA extraction, fluorescence PCR reaction detection and dissolving curve analysis. The detection kit is operated simply and quickly, intuitive in result, high in sensitivity, high in specificity and high in stability and can be widely applied to quick identification of the legionella pneumophila in clinical and environment water samples.

Description

technical field [0001] The invention belongs to the field of medicine, in particular to a kit for detecting Legionella pneumophila by real-time fluorescent PCR melting curve. Background technique [0002] Legionella, a facultative intracellular parasite, is an important pathogen that causes legionnaires' disease in humans. In recent years, Legionnaires' disease has broken out frequently around the world, and my country has listed it as one of the 14 emerging infectious diseases. According to foreign reports, 10% of nosocomial pneumonia infections of unknown cause are caused by Legionella. Although currently known Legionella has 52 species, more than 70 serotypes. But clinically more than 80% of Legionella infections are caused by Legionella pneumophila. Therefore, the identification of Legionella pneumophila is of great value in the diagnosis of Legionnaires' disease. [0003] At present, the detection and identification methods of Legionella mainly include isolation and...

AI Technical Summary

Problems solved by technology

However, the above detection methods all have deficiencies, such as direct isolation and culture, the time required is as long as 3 to 10 days, and the positive rate is low; urine antigen detection is only specific to Legionella pneumophila type 1; molecular techniques also have their own limitations. Disadvantages, such as traditional polymerase chain reaction (PCR) detection is cumbersome, time-consuming, and easy to contaminate; hybridization probe labeling technology based on the principle of fluorescence resonance energy transfer is expensive and complicated to operate; DNA sequencing is the most accurate identification of Legionella species method, but it takes a long time, analysis is difficult, and cannot meet the rapid clinical requirements, and other molecular detection methods have various deficiencies

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