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Primer group, kit and method for detecting respiratory pathogens

A pathogen and respiratory technology, applied in the field of respiratory pathogen detection, can solve the problems of large impact on results, inability to distinguish contamination and colonization bacteria, long time consumption, etc., to reduce the cost of detection and avoid limited detection channels.

Inactive Publication Date: 2019-03-12
SUZHOU INST OF NANO TECH & NANO BIONICS CHINESE ACEDEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this method takes a long time, has low sensitivity, and requires high experimental operations; in addition, there are bacterial culture and drug sensitivity methods, which can detect drug resistance, but the samples are easily contaminated, which has a great impact on the results; the determination of viral antigens can be used as evidence of infection , the detection is fast, but the sensitivity is low, and it cannot distinguish between pollution and colonized bacteria; although antibody detection is easy to operate, its sensitivity and specificity vary greatly, and false positive and false negative results are prone to occur

Method used

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  • Primer group, kit and method for detecting respiratory pathogens
  • Primer group, kit and method for detecting respiratory pathogens
  • Primer group, kit and method for detecting respiratory pathogens

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] The primer set for detecting respiratory pathogens provided in this embodiment includes the first primer pair-the thirteenth primer;

[0071] Wherein, the first primer pair includes the primers shown in SEQ ID NO.1~2, the second primer pair includes the primers shown in SEQ ID NO.3~4, and the third primer pair includes the primers shown in SEQ ID NO.5~6. primers, the fourth primer pair includes the primers shown in SEQ ID NO.7-8, the fifth primer pair includes the primers shown in SEQ ID NO.9-10, and the sixth primer pair includes the primers shown in SEQ ID NO.11-12 The primers shown, the 7th primer pair includes the primers shown in SEQ ID NO.13-14, the 8th primer pair includes the primers shown in SEQ ID NO.15-16, the 9th primer pair includes the primers shown in SEQ ID NO. The primers shown, the 10th primer pair includes the primers shown in SEQ ID NO.19-20, the 11th primer pair includes the primers shown in SEQ ID NO.21-22, and the 12th primer pair includes the pri...

experiment example 1

[0097] (1) Specificity verification

[0098] Use each respiratory pathogen plasmid standard to do a cross experiment to verify its specificity

[0099] Taking RSV as an example, add the same primer pair to the wells of the same row of PCR 8-tube tubes (only 1 pair of primers is added to each tube, a total of 13 rows, and rows 1-13 correspond to the added primer pair respectively as the first primer Pair - the 13th primer pair), the same RSV positive plasmid (ie, the PUC57 plasmid containing the RSV conservative sequence) template was added to the same row of wells to ensure that each primer pair had independent contact with any of the 13 respiratory pathogens Opportunity, through such a crossover experiment, count each CT value, and verify the specificity of the primers.

[0100] The reaction solution configuration method is as follows: 1X PCR reaction master mix containing SYBR Green I; 300nM upstream primer; 300nM downstream primer; 0.2unit uracil-DNA glycosylase (UNG enzym...

experiment example 2

[0112] Experimental example 2 uses other control primers to carry out the crossover experiment as shown in Table 6 below: Since SYBR is a non-specific dye, these control primers show more non-specific amplification in the crossover experiment. This shows that the first primer pair-the thirteenth primer pair of the present invention is used to detect respiratory pathogens such as respiratory syncytial virus, adenovirus, human metapneumovirus, rhino / enterovirus, parainfluenza virus, coronavirus, boca virus , Influenza virus A, Influenza virus B, B. pertussis, B. parapertussis, Mycoplasma pneumoniae and Chlamydia pneumoniae have high specificity.

[0113] The sequence of the control primer corresponding to each virus is supplemented in turn:

[0114]

[0115] Table 6

[0116]

[0117] It can be seen that the first primer pair-the thirteenth primer pair of the present invention are used to detect respiratory pathogens such as respiratory syncytial virus, adenovirus, human m...

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Abstract

The invention discloses a primer group, a kit and a method for detecting respiratory pathogens and relates to the technical field of respiratory pathogen detection. The primer group for detecting therespiratory pathogens comprises, such as, one or a combination of several of a first primer to a thirteenth primer. The primer group can realize detection of common respiratory pathogens such as a respiratory syncytial virus, an adenovirus, a human metapneumovirus, a rhinovirus / enterovirus, a parainfluenza virus, a coronavirus, a bocavirus, an influenza virus A, an influenza virus B, bordetella pertussis, bordetella parapertussis, mycoplasma pneumoniae and chlamydia pneumoniae and has the characteristics of relatively high sensitivity and specificity, low cost, convenience in detection, relatively good repeatability and the like.

Description

technical field [0001] The invention relates to the technical field of detection of respiratory pathogens, in particular to a primer set, a kit and a method for detecting respiratory pathogens. Background technique [0002] Acute respiratory infections (ARIs) are one of the most common infectious diseases in humans worldwide and one of the important causes of morbidity and mortality worldwide. Acute respiratory tract infection is more common in children, the elderly and immunocompromised persons, especially common and frequently-occurring diseases in pediatrics. [0003] Acute respiratory tract infections can be divided into upper respiratory tract infections (URIs) and lower respiratory tract infections (LRIs). The most common symptoms of upper respiratory tract infection include sore throat, nasal congestion, headache, sneezing, coughing, and fever. About 90% of them are caused by viruses. The most common symptoms of lower respiratory tract infection are similar to those...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/689C12Q1/686C12Q1/04C12N15/11
CPCC12Q1/686C12Q1/689C12Q1/701C12Q2600/16C12Q2600/166C12Q2537/143C12Q2545/113C12Q2563/107Y02A50/30
Inventor 李炯王娜娜刘胜权
Owner SUZHOU INST OF NANO TECH & NANO BIONICS CHINESE ACEDEMY OF SCI
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