Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Real-time fluorescent quantitative PCR detection primer, probe, kit and method for Ljungan virus and application thereof

A technology for real-time fluorescence quantification and primer detection, which is applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc. Contamination, the effect of shortening the operation time

Pending Publication Date: 2020-01-17
广东龙帆生物科技有限公司
View PDF3 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, the current detection methods for Yonganhe virus are mainly traditional electron microscope observation and serum neutralization. These detection methods have disadvantages such as cumbersome operation, long cycle, and poor specificity. How to identify viruses

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 fluorescent quantitative PCR detection primer, probe, kit and method for Ljungan virus and application thereof
  • Real-time fluorescent quantitative PCR detection primer, probe, kit and method for Ljungan virus and application thereof
  • Real-time fluorescent quantitative PCR detection primer, probe, kit and method for Ljungan virus and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] The development of embodiment 1 real-time fluorescent quantitative PCR detection kit

[0033] 1. In this embodiment, select the same or similar segment of the specific VP1 gene sequence of Yonganhe virus to design a pair of specific primers and a specific fluorescent probe, select the internal standard gene Rnase P to design a pair of specific Primers and a specific fluorescent probe can be used to detect Yonganhe virus by constructing real-time fluorescent quantitative PCR technology. The VP1 gene is the antigenic determinant gene of Yong'anhe virus, and the species can be determined based on the VP1 sequence, while other techniques generally design primers and probes for the 5'UTR region of the virus, which cannot determine the virus species. The primer sequences and probe sequences designed in this embodiment of the present invention are shown in Table 1 below.

[0034] Table 1

[0035]

[0036] The kit also includes positive control: Yonganhe virus standard; ne...

Embodiment 2

[0070] The performance measurement of the real-time fluorescent PCR kit of embodiment 2 Yong'an River virus

[0071] 1. Verification of accuracy

[0072] The gold standard for viral nucleic acid detection is genome sequencing. The test results of this kit are compared with viral genome sequencing to analyze its accuracy. In this example, 22 samples of Yonganhe virus were selected by genome sequencing, including 22 samples of Yonganhe virus. The results of detection with the kit provided by the present invention are shown in Table 4 below. It can be seen from the results that all 22 positive specimens were detected, indicating that the accuracy of the real-time fluorescent PCR of Yonganhe virus provided by the present invention is 100%.

[0073] Accuracy analysis of the present invention of table 4

[0074] virus type Number of cases Sequencing positive This method is positive Accuracy Yongan River virus 22 22 22 100％

[0075] 2. Specificity Ve...

Embodiment 3

[0090] Embodiment 3 clinical detection

[0091] The above method was used to test the stool samples of 20 other patients with suspected Yonganhe virus infection, among which 3 cases were positive for Yonganhe virus virus detection, the virus fluorescence quantitative PCR amplification curve is shown in Image 6 , according to the C of these 3 positive results t Combined with the amplification curve, Roche LightCycler 480 analysis software automatically analyzed the virus concentration of these 3 cases of Yonganhe virus-positive samples. The specific results are shown in Table 8. At the same time, the amplification curves of the internal reference gene Rnase P in these 20 cases of stool samples were normal, as shown in Figure 7 As shown, it shows that the extraction and amplification process of this experiment is normal, and the positive and negative results are accurate.

[0092] Table 8 Virus concentration of 5 cases of positive specimens of Yonganhe virus

[0093] ...

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

PropertyMeasurementUnit
Sensitivityaaaaaaaaaa
Login to View More

Abstract

The invention belongs to the technical field of virus detection, and particularly relates to a real-time fluorescent quantitative PCR detection primer, probe, kit and method for Ljungan virus and application thereof. Two single-tube fluorescence channels are adopted to simultaneously detect the existence of the Ljungan virus and reference gene Rnase P, and can detect the existence of the RNA of the Ljungan virus in specimens such as whole blood, serum, cerebrospinal fluid, excrement and tissues. According to the invention, the detection time period is short and the detection efficiency is high; the virus detection specificity is high, and the accuracy is high; virus quantitative analysis can be carried out while virus qualitative analysis is carried out, and the quantitative linear range is good; the detection sensitivity is high; operation is simple, and popularization is easy; experimental results are good in repeatability and high in precision; and the quality of the whole process of extracting and amplifying the specimens can be monitored through the detection result of the reference gene.

Description

technical field [0001] The invention belongs to the technical field of virus detection, and in particular relates to a real-time fluorescent quantitative PCR detection primer and probe for Yonganhe virus, a detection kit, a detection method and an application thereof. Background technique [0002] Ljungan virus (Ljungan virus) belongs to Picornaviridae and Paretrovirus genus. It is a zoonotic virus discovered in 1999. It can transmit infection between humans and rodents. Ljungan virus infects It can lead to fetal malformation, fetal death, sudden neonatal death and myocarditis, etc. It is also found that Yonganhe virus infection has a significant correlation with diabetes. [0003] However, the current detection methods for Yonganhe virus are mainly traditional electron microscope observation and serum neutralization. These detection methods have disadvantages such as cumbersome operation, long cycle, and poor specificity. Methods for the identification of viruses. Conten...

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): C12Q1/70C12Q1/6851C12N15/11
CPCC12Q1/701C12Q1/6851C12Q2600/166C12Q2531/113C12Q2563/107C12Q2545/101C12Q2545/113C12Q2547/101
Inventor 吴建国刘为勇刘映乐谭秋萍
Owner 广东龙帆生物科技有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com