Duplex PCR diagnostic kit for detection of contagious caprine pleuropneumonia pathogen and preparation and use methods thereof

Abstract

The invention discloses a raw material composition of a duplex PCR (polymerase chain reaction) diagnostic kit for detection of contagious caprine pleuropneumonia pathogen and a preparation method and a use method of the kit. The raw material composition comprises 1000-2000muL of PCR reaction liquid, 100-200muL of TaqDNA polymerase, 50-100muL of positive control, 50-100muL of negative control and 1-2mL of ultrapure water. The PCR reaction solution comprises a primer 1 mixture solution, a primer 2 mixture solution, a PCR reaction buffer and dNTPs (deoxyribonucleotide triphosphates), wherein the volume ratio of the four liquids in the PCR reaction solution is 2:2:5:1. The preparation method comprises the steps of determination of an optimum reaction annealing temperature, specific detection, sensibility test, clinical application detection and kit packaging. An effective method for rapid deterministic diagnosis of suspected goat pathogen and suspected cases of contagious caprine pleuropneumonia in goat farms is provided by the invention, providing technical support for the prevention and control of contagious caprine pleuropneumonia.

Description

technical field [0001] The invention relates to a goat infectious pleuropneumonia pathogen detection double PCR diagnostic kit and a preparation method. Background technique [0002] Contagious Caprine Plneuropneumonia (CCPP), also known as rotten lung disease, is a highly contagious infectious disease of goats. The Ministry of Agriculture of my country lists it as a second-class infectious disease. It clinically presents with fibrous pneumonia and pleurisy. The mortality rate is higher. Mycoplasma mycoides cluster members Mycoplasma capricolum subsp. capripneumoniae (Mccp), Mycoplasma mycoides subsp. mycoides LC (Mmm LC) and Mycoplasma mycoides subsp. Mycoplasma mycoides subsp.capri (Mmc) can cause goats to suffer from CCPP, and Mycoplasma ovipneumoniae (Mo) can also infect goats. [0003] At present, the diagnosis of CCPP at home and abroad mainly relies on pathogen isolation and identification, serological tests and molecular biology techniques. Compared with the three ...

AI Technical Summary

Problems solved by technology

Compared with the three diagnostic methods, the traditional separation and identification of mycoplasma has high requirements on growth conditions and slow growth rate, so the diagnosis cost is high, difficult and time-consuming, and cannot immediately solve the problem of rapid diagnosis; serological tests are mainly indirect hemagglutination Test, complement fixation test and competitive ELISA, the three have poor sensitivity or specificity, and can not be used as a pathogenic diagnosis method; molecular biology technology has the advantages of fast, accurate, sensitive, specific, etc., especially the multiplex PCR method can be used for different pathogens at the same time diagnosis

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