Primer probe group and kit for combined detection of mycoplasma pneumoniae and chlamydia pneumoniae based on fluorescence RMA method

Abstract

The invention belongs to the technical field of detection of mycoplasma pneumoniae and chlamydia pneumoniae, and particularly relates to a primer probe group, a kit and a detection method for combineddetection of mycoplasma pneumoniae and chlamydia pneumoniae based on a fluorescence RMA method. The kit comprises a detection tube containing an amplification reaction reagent, a buffer solution, magnesium acetate, standard positive plasmids and sterile double distilled water. The amplification reaction reagent is prepared from primers and probes of MP and Cpn, M-MLV reverse transcriptase, escherichia coli RecA protein, UvsY protein, single-stranded binding protein GP32, Bst polymerase, exonuclease III, polyethylene oxide, trehalose, mannitol, ATP, dNTPs, creatine kinase and creatine phosphate. The standard positive plasmids are recombinant plasmids containing MP and Cpn amplified gene sequences, and are used for positive control of MP and Cpn nucleic acid detection.

Description

technical field [0001] The application belongs to the technical field of detection of Mycoplasma pneumoniae and Chlamydia pneumoniae, and specifically relates to a primer-probe set, a kit and a detection method for combined detection of Mycoplasma pneumoniae and Chlamydia pneumoniae based on a fluorescent RMA method. Background technique [0002] Mycoplasma pneumoniae (MP) is one of the important pathogens of community-acquired pneumonia (CAP) in children. Mycoplasma pneumoniae is a microorganism between bacteria and viruses. It is mainly transmitted through the respiratory tract. Mycoplasma can spread to any organ in the body through blood. In addition, mycoplasma infection can produce autoantibodies, resulting in multi-system immune damage, causing multiple The system is malfunctioning. Mycoplasma pneumoniae pneumonia (MPP) accounts for 10%-40% of CAP in hospitalized children and is a widespread concern for pediatricians. [0003] Chlamydia pneumoniae (Cpn) is an obligat...

AI Technical Summary

Problems solved by technology

[0004] At present, the commonly used detection methods for Mycoplasma pneumoniae and Chlamydia pneumoniae are as follows: (1) Virus isolation and culture method: It is the most reliable method for diagnosing MP and Cpn infection, but so far, these two pathogens are difficult to obtain through culture, On the one hand, because most of these patients have dry cough, it is difficult to obtain sputum samples; on the other hand, the culture requirements for mycoplasma and chlamydia are very high, and it is difficult for general laboratories

(2) Serological detection: The most commonly used serological method is complement fixation reaction (CFTs), but there are many shortcomings in this method, such as the false positive of mycoplasma CFTs can appear in the inflammatory reaction process, and CFTs cannot be used to distinguish Chlamydia pneumoniae from Chlamydia psittaci infection, because both have a common antigen; enzyme-linked immunosorbent assay (ELISA) can distinguish acute infection from past infection because it can detect IgM and IgG in serum, and is suitable for clinical laboratories as a routine for MP and Cpn infection Check, the disadvantage is poor sensitivity, low positive predictive value, not suitable for crowd screening

(3) Molecular biology detection: the detection of MP and Cpn nucleic acid in clinical specimens by molecular biology techniques has high sensitivity and specificity, but attention must be paid to quality control, otherwise more false positive results may occur, and Requires special PCR amplification conditions, specialized laboratories and PCR machines, and is prone to contamination during the detection process

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