Real-time fluorescence multiplex PCR primer probe for seven common respiratory system influenza virus pathogens and kit

A technology of influenza virus and primer probe, which is applied in the biological field to achieve the effect of saving time and preventing public health emergencies

Active Publication Date: 2018-04-20
北京卓诚惠生生物科技股份有限公司
View PDF3 Cites 15 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the problem of simultaneous detection and identification of multiple respiratory viruses, provide a fluorescent quantitative PCR primer probe set

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Real-time fluorescence multiplex PCR primer probe for seven common respiratory system influenza virus pathogens and kit
  • Real-time fluorescence multiplex PCR primer probe for seven common respiratory system influenza virus pathogens and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0116] Formation of the first system:

[0117] Primer probes include:

[0118] The parainfluenza virus type I upstream primer of the nucleotide sequence shown in SEQ ID NO.1,

[0119] The parainfluenza virus type I downstream primer of the nucleotide sequence shown in SEQ ID NO.2,

[0120] A parainfluenza virus type I probe having the nucleotide sequence shown in SEQ ID NO.3, the 5' end of the parainfluenza virus type I probe is marked with a CY5 luminescent group, and the 3' end is marked with a fluorescent quencher Mission BHQ3;

[0121] The parainfluenza virus type Ⅱ upstream primer of the nucleotide sequence shown in SEQ ID NO.4,

[0122] Parainfluenza virus type Ⅱ downstream primer of the nucleotide sequence shown in SEQ ID NO.5,

[0123] A parainfluenza virus type II probe having the nucleotide sequence shown in SEQ ID NO.6, the 5' end of the parainfluenza virus type II probe is marked with a VIC luminescent group, and the 3' end is marked with a fluorescent quencher...

Embodiment 2

[0145] Operation and result judgment of embodiment 2 kit

[0146] 1. Genome extraction

[0147] After the throat swab is put into the sampling solution, it needs to be fully shaken and washed, and then left to stand at room temperature for 10 minutes. After the large lumps sink, the supernatant is taken and centrifuged immediately, and the subsequent precipitation can be used for nucleic acid extraction.

[0148] 2. Preparation of reaction system

[0149] The reaction system detected by the kit is 25 μL, and its configuration is as follows: 5 μL of 5× RT-PCR buffer (the buffer in tube B contains IAC template); 2.5 μL of 10× enzyme mixture; 2.5 μL of 10× primer-probe mixture ( The concentration of each primer is 0.5 μM, the concentration of each probe is 0.2 μM), RNA template 5 μL, and ultrapure water to make up to 25 μL.

[0150] 3. PCR reaction

[0151] Put the PCR tube into the fluorescence quantitative PCR instrument, and carry out the PCR reaction according to the follo...

Embodiment 3

[0157] The shelf life test of embodiment 3 kit

[0158] respectively by 10 5 PFU / mL The mixed template of parainfluenza virus type I, parainfluenza virus type II, parainfluenza virus type III and influenza B virus and the mixed template of adenovirus, respiratory syncytial virus and influenza A virus were used as test samples for evaluation, On day 0, aliquots were divided into 9 portions and stored in a -70°C refrigerator. The completed kits were stored at -20°C, and the kits of 0, 10, 15, 30, 60, 90, 120, 150, 180 and 200 days were used for storage test. The test results of the shelf life are shown in Table 2:

[0159] Table 2 shelf life test results

[0160] shelf life

[0161] It can be seen from Table 2 that the kit is stored in a -20°C refrigerator, and it is effective in detecting the seven target viruses at different storage periods. The experimental results show that the storage period of the kit is at least 6 months.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The present disclosure relates to a real-time fluorescence multiplex PCR primer probe for seven common respiratory system influenza virus pathogens and a kit. Seven influenza viruses are influenza A virus, influenza B virus, parainfluenza virus type I, parainfluenza virus type II, parainfluenza virus type III, respiratory syncytial virus and adenovirus. The present disclosure also provides a kit for multiplex fluorescence quantitative PCR detection of the seven influenza viruses, and the kit includes the primer probe set. The kit significantly improves the sensitivity, specificity, and simplicity of detection of the common respiratory system pathogens.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a real-time fluorescent multiplex PCR primer probe and a rapid detection kit for seven common influenza virus pathogens in the respiratory system. Background technique [0002] Respiratory tract infections are common among adults and children and are associated with high morbidity and mortality. Acute upper and lower respiratory infections are mainly spread through droplets in the air, person-to-person contact or contact with contaminated items. Clinically common respiratory viruses include influenza A virus, influenza B virus, respiratory syncytial virus, adenovirus, parainfluenza type I virus, parainfluenza type II virus and parainfluenza type III virus. Because the infection symptoms and epidemic characteristics caused by these viruses are similar, it is very unreliable to distinguish the pathogens only by clinical symptoms. Laboratory diagnosis methods include virus isolation, a...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11
CPCC12Q1/6851C12Q1/701C12Q2600/16C12Q2537/143C12Q2563/107C12Q2531/113
Inventor 林笑冬凌明玉王雷张志强
Owner 北京卓诚惠生生物科技股份有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap