Real-time fluorescence quantification PCR detecting kit for cow mycoplasma and special primers and TaqMan probe thereof

Abstract

The invention discloses a real-time fluorescence quantification PCR detecting kit for cow mycoplasma, special primers and a TaqMan probe thereof and application of the detecting kit in detection of cow mycoplasma. The primers and the TaqMan probe for real-time fluorescence quantification PCR detecting of the cow mycoplasma are designed according to a specific conserved sequence of an OPPD / F gene of the cow mycoplasma and are used for detecting the cow mycoplasma of a sample to be detected qualitatively and quantitatively. The nucleotide sequence of the upstream primer (OF-A) is shown as SED ID NO:1 in a sequence table, the nucleotide sequence of the downstream primer (OF-B) is shown as SED ID NO:2 in the sequence table, and the nucleotide sequence of the TaqMan probe (OF-P) is shown as SEQ IDNO:3 in the sequence table. By means of the detecting kit, the special primers and the TaqMan probe thereof, the cow mycoplasma can be detected quickly, conveniently, efficiently, highly specifically and highly sensitively, and a novel technical platform is provided for detection of the cow mycoplasma.

Description

Technical field [0001] The invention belongs to the detection of bovine mycoplasma in the field of veterinary infectious diseases, and particularly relates to a real-time fluorescent quantitative PCR detection kit for detecting bovine mycoplasma pathogens, its special primers, TaqMan probes and their application in detecting bovine mycoplasma. Background technique [0002] Mycoplasma bovis (Mycoplasmabovis) belongs to the class of Lapis, Mycoplasma, Mycoplasma, and Mycoplasma. It is the smallest microorganism known to grow and reproduce independently in a cell-free medium. In 1961, American scientist Hale isolated Mycoplasma bovis from cow's milk suffering from mastitis for the first time. It was first reported in 1976 that Mycoplasma bovis was related to bovine respiratory diseases. In 2008, mycoplasma bovis, which can cause pneumonia, was isolated and identified for the first time in my country, and the pathogen of this disease was successively isolated in Hubei, Inner Mongoli...

AI Technical Summary

Problems solved by technology

Etiological detection is mainly the isolation and culture of pathogenic microorganisms and the staining microscope examination of pathogenic microorganisms. Since Mycoplasma bovis has strict requirements on nutrients and a long culture period, it is pleomorphic under the microscope and has a low detection rate. For the clinical diagnosis of bovis mycoplasma disease, routine pathogenic detection has too many disadvantages

The most widely used immunological detection method is enzyme-linked immunosorbent assay (Enzyme-linkedimmunosorbentassay, ELISA). Most ELISA kits are coated with whole bacterial protein as antigen. The surface membrane proteins are very similar, and it is still difficult to find a highly specific antigen for differential diagnosis at this stage, and the ELISA diagnostic method coated with whole bacterial protein as an antigen is bound to have cross-reactivity, resulting in poor specificity of the method

The increasing development of molecular biology technology is of great significance for the clinical diagnosis of infectious diseases. At present, the molecular biology detection methods for Mycoplasma bovis are widely used in China, mainly including multiplex PCR (multiplexPCR), nested PCR (nestedPCR), RT- PCR (reversetranscriptasePCR) etc., and the real-time fluorescent quantitative PCR detection method of Mycoplasma bovis has not been reported yet

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