Influenza A virus and B virus joint detection primer, probe, kit and application

An influenza virus, combined detection technology, applied in the field of kits and applications, B type combined detection primers, influenza virus A type, probe fields, can solve the problem of reducing operation steps, etc., to reduce operation steps, improve detection efficiency, improve Detecting the effect of speed

Inactive Publication Date: 2015-04-29
深圳澳东检验检测科技有限公司
View PDF3 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The invention is used in RT-PCR, which overcomes the disadvantage of traditional RT-PCR that needs to extract RNA, which is a cumbersome step, reduces operating steps, can detect influenza virus types A and B at the same time, improves detection efficiency, saves costs, and effectively avoids crossover High-throughput detection of influenza virus is of great significance for rapid detection of influenza epidemics and timely formulation of prevention and control measures

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
  • Influenza A virus and B virus joint detection primer, probe, kit and application
  • Influenza A virus and B virus joint detection primer, probe, kit and application
  • Influenza A virus and B virus joint detection primer, probe, kit and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 2

[0068] Embodiment 2 tests the sensitivity of the detection method of embodiment 1

[0069] Using the optimal reaction system in step 2 and the optimal reaction conditions in step 3, perform direct amplification RT-PCR on throat swab samples and allantoic fluid samples, and obtain the amplification curve and Ct of each reaction tube from the fluorescent quantitative PCR. Values, the average Ct of each sample was calculated. From figure 1 It can be seen that for the throat swab sample, using 1 μL of purified AIV RNA as a template, the Ct value of the fluorescent quantitative PCR was 19.42 for the A-type nucleic acid sample, and 19.53 for the B-type nucleic acid sample, while the direct amplification RT of Example 1 was used. -The Ct value detected by PCR from 1 μL AIV PBS eluate is 18.93 for type A nucleic acid samples, and 21.23 for type B nucleic acid samples (such as figure 1 ).

[0070] For allantoic fluid samples, using 1 μL of purified AIV RNA as a template, the Ct valu...

Embodiment 3

[0071] Embodiment 3 tests the tolerance of throat swab fluid of the detection method of embodiment 1

[0072]According to the optimal reaction system in step 2 of Example 1, the reaction solution except the throat swab solution was prepared; 1 μL AIV positive throat swab solution was diluted to 2.5, 5, 7.5, respectively with the negative throat swab solution not containing AIV. 10 μL were added to the reaction tubes to keep the AIV virus copy numbers in each tube consistent, and the percentages of throat swab fluid in the total volume were 10%, 20%, 30%, and 40%, respectively. Put each reaction tube into a fluorescent quantitative PCR instrument, and perform the reaction according to the optimal reaction conditions in Step 3 of Example 1.

[0073] The result is as image 3 As shown, the direct amplification RT-PCR still has high activity in up to 40% throat swab fluid, and the change of Ct value is less than 2 values ​​compared with 10% concentration, which has high tolerance...

Embodiment 4

[0074] Embodiment 4 Test the detection dynamic range and the linearity of the detection method of embodiment 1

[0075] According to the optimal reaction system in step 2 of Example 1 to prepare the reaction solution except for the sample; take a positive sample of AIV throat swab and dilute it with a negative throat swab that does not contain AIV to obtain a 10-fold gradient of positive Throat swab fluid sample 10 0 ,10 -1 ,10 -2 ,10 -3 , all reaction tubes maintain a final concentration of 5% of the throat swab fluid. Take 10 μL of throat swab liquid of various dilutions and add them to reaction tubes, place them in a fluorescent quantitative PCR instrument, and perform the reaction according to the optimal reaction conditions in Step 3 of Example 1.

[0076] The results showed that the pharyngeal swab liquid of 1000 times dilution still obtained positive detection signal ( Figure 4 Fluorescent PCR curves for each concentration, Figure 5 is a linear range graph); the...

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 invention discloses an influenza A virus and B virus joint detection primer, a probe, a kit and an application. The sequence of the primer is as shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 4 and SEQ ID NO: 5; and the sequence of the probe is as shown in SEQ ID NO: 3 and SEQ ID NO: 6. The primer and the probe disclosed by the invention, when used for detecting reverse transcription-polymerase chain reaction (RT-PCR) which is directly amplified by influenza A virus and B virus, can be used for overcoming the shortcomings of a complex step of extracting RNA from conventional RT-PCR and reducing operation steps; the influenza A virus and B virus can be simultaneously detected, so that detection efficiency is improved, cost is saved and cross pollution is effectively avoided; the primer and the probe can be used for the high-throughput detection of influenza viruses, and is of great significance for the fast discovery of influenza epidemic situation and for the timely formulation of prevention and control measures.

Description

technical field [0001] The invention relates to the technical field of molecular biology detection of influenza virus, in particular to primers, probes, kits and applications for combined detection of influenza virus A and B. Background technique [0002] Influenza virus (influenza virus), referred to as influenza virus, includes human influenza virus and animal influenza virus. Human influenza virus is divided into three types: A (A), B (B), and C (C). ) pathogens. The antigenicity of the nucleoprotein of influenza virus infecting birds, pigs and other animals is the same as that of human influenza A virus. [0003] In recent years, it has been found that influenza not only seriously harms poultry, pigs and other animals, but especially can infect and cause death in humans, which greatly threatens the safety of human public health. The genetic material of influenza virus consists of eight independent single-stranded negative-sense RNA segments of varying sizes. The seven...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
CPCC12Q1/701C12Q1/686C12Q2600/112
Inventor 翟建新康康潘艳萍苟德明张利
Owner 深圳澳东检验检测科技有限公司
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