Triple fluorogenic quantitative PCR detection method of escherichia coli, Klebsiella pneumoniae and acinetobacter

Abstract

The invention relates to a triple fluorogenic quantitative PCR detection method of escherichia coli, Klebsiella pneumoniae and acinetobacter, and belongs to the technical field of fluorogenic quantitative PCR detection. The detection method comprises the following steps: preparing a 25ul reaction system by 10ul nucleic acid of a sample and a 15ul reaction liquid, and then carrying out fluorogenic quantitative PCR reaction; and determining that the sample contains escherichia coli if an S-shaped amplification curve is existent in the FAM detection channel and Ct value is less than 30, determining that the sample contains Klebsiella pneumoniae if the S-shaped amplification curve is existent in the HEX or VIC detection channel and Ct value is less than 30, and determining that the sample contains acinetobacter if the S-shaped amplification curve is existent in CY5 detection channel and Ct value is less than 30. The whole detection process is less than two hours, a closed-tube reaction is adopted, and the reaction system is processed directly after reaction, so the aerosol pollution caused by opening is avoided. The process is simple and the detection time is short.

Description

technical field [0001] The invention relates to a triple fluorescent quantitative PCR detection method for Escherichia coli, Klebsiella pneumonia and Acinetobacter, belonging to the technical field of fluorescent quantitative PCR. Background technique [0002] In recent years, relying on its unique climate characteristics and high-quality and rich feed resources, Shandong Province has become the largest province in the country for fur animal breeding, and the fur animal breeding accounts for more than 55% of the total national breeding. Among them, the breeding volume of mink accounts for more than 80% of the country's total, and the breeding volume of blue fox accounts for more than 50% of the country's total. The economic benefits are considerable, and it has become a characteristic animal husbandry in Shandong Province. However, with the vigorous development of the fur animal breeding industry, the contradiction between industrial development and resource constraints has ...

AI Technical Summary

Problems solved by technology

At the same time, 16sRNA identification also needs to be sequenced, and the cost is relatively high

Ordinary PCR identification has poor sensitivity, and sometimes bacteria cannot be detected directly from the sample, and it is easy to contaminate and has high false positives

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