Creation of sheep-derived pasteurella multocida fluorescent quantitative PCR (polymerase chain reaction) standard curves

Abstract

The invention discloses creation of sheep-derived pasteurella multocida fluorescent quantitative PCR (polymerase chain reaction) standard curves. The creation has the advantages that toxA toxin genesin Pasteurella multocida genomes published in GenBank are taken as reference, conservative sequences cstoxA are acquired by the aid of information analysis software, recombinant plasmids pMD-19T-cstoxA of positive standard substances are constructed by means of PCR, fluorescent quantitative PCR systems are optimized, accordingly, the sheep-derived pasteurella multocida fluorescent quantitative PCRstandard curves can be created, pasteurella multocida fluorescent quantitative PCR standard curves with high specificity and sensitivity and good repeatability can be obtained, convenient, speedy andaccurate bacterial strain typing effects and the like can be realized, the specificity can be improved as compared with the traditional laboratory PCR operation, and the expenditure for laboratory consumables and the like further can be reduced as compared with the traditional laboratory PCR operation.

Description

technical field [0001] The invention belongs to the field of fluorescent quantitative PCR, and in particular relates to the establishment of a fluorescent quantitative PCR standard curve of sheep-derived Pasteurella multocida. Background technique [0002] Pasteurellosis, also known as hemorrhagic sepsis, is a zoonotic infectious disease characterized by sepsis and pneumonia caused by Pasteurella multocida (Pm). Its serotypes are diverse and its pathogenicity is complex. Pasteurella multocida can be typed by serotyping method, and can be divided into five serotypes A, B, D, E, and F according to different capsular antigens (K antigen), and according to lipopolysaccharide antigen (O Antigens) can be divided into 16 serotypes (1-16). Clinically, due to differences in antigenicity and host specificity, various clinical symptoms have been caused, such as atrophic rhinitis in pigs, hemorrhagic sepsis in cattle and sheep, and fowl cholera in poultry. In summary, a rapid and eff...

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Problems solved by technology

With the continuous in-depth research on PCR technology, the traditional PCR method requires electrophoresis analysis, which is easy to cause pollution and takes a long time. It can only be qualitative but not quantitative.

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