RPA detection method, special primers and probe for detecting Rickettsia rickettsii, and uses of RPA detection method in detection of Rickettsia rickettsii

Abstract

The invention provides an RPA detection method, special primers and a probe for detecting Rickettsia rickettsii, and uses of the RPA detection method in detection of Rickettsia rickettsii. According to the present invention, the special primers and the probe are designed based on the conserved sequence of a Rickettsia rickettsii gene, and have the oligonucleotide sequences represented by SEQ ID NO.2, SEQ ID NO.3 and SEQ ID NO.4; the novel constant temperature amplification technology RPA is firstly applied in the detection of Rickettsia rickettsii, wherein the enzymatic reaction process of DNAreplication in vivo is simulated, the DNA template is amplified by using the specific enzyme and protein combination including recombinase, single-stranded binding protein and DNA polymerase so as toamplify the specific nucleic acid sequence at the constant temperature of 37 DEG C, and the amplification product can be subjected to visualized discrimination through the lateral chromatography nucleic acid detection test paper strip; and the established detection method has the sensitivity of 6 copies / [mu]L, has high specificity, has low requirement on hardware equipment, can complete the detection within 30 min, does not require the complicated sample treatment, is suitable for on-site detection, and is suitable for popularization and application.

Description

technical field

[0001] The invention belongs to the field of biotechnology, relates to the molecular biology of Rickettsia rickettsia, relates to a method for detecting Rickettsia rickettsia and its application, and particularly relates to a technique of using recombinase polymerase constant temperature amplification (RPA Technology) rapid detection method for Rickettsia rickettsiae, its special primers and probes and its application. Background technique

[0002] Rickettsia rickettsiae ( Rickettsia rickettsii , Rr), commonly known as spotted fever, is the pathogen of Rocky Mountain spotted fever. It is a small obligate intracellular gram-negative bacterium that has strong resistance to the external environment and a long survival time. Humans and animals are generally susceptible, and can pass through Aerosols are widespread and are among the most lethal, biologically agentized pathogens. Rickettsia rickettsii mainly invades arteriovenous endothelial cells, causing vascul...

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