Reverse transcription loop-mediated isothermal amplification (RT-LAMP) kit for porcine kobuvirus and application thereof

A loop-mediated isothermal, crista virus technology, applied in the field of microbial detection, can solve the problems of difficulty in detecting porcine crista virus, time-consuming, expensive instruments, etc., and achieve the effect of simple interpretation method, simple operation, and rapid acquisition.

Inactive Publication Date: 2016-08-03
广西农垦永新畜牧集团金光有限公司 +1
View PDF2 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to solve the problems such as difficulty in detection of porcine crest virus at the grassroots level, time-consuming and expensive equipment, and provide a simple, fast and accurate detection kit for the basic level of porcine crest virus, the technical solution used to achieve the purpose of the present invention It is: a porcine ridge virus reverse tra

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
  • Reverse transcription loop-mediated isothermal amplification (RT-LAMP) kit for porcine kobuvirus and application thereof
  • Reverse transcription loop-mediated isothermal amplification (RT-LAMP) kit for porcine kobuvirus and application thereof
  • Reverse transcription loop-mediated isothermal amplification (RT-LAMP) kit for porcine kobuvirus and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] The specificity result of embodiment 1RT-LAMP detection method

[0082] RT-LAMP amplification was performed on 1 strain of porcine ridge virus, 7 strains of control virus and water control, and the results were as follows figure 1 As shown, the porcine crest virus reaction tube showed a rising curve of turbidity at about 23 minutes, which was a positive result, and the 7 strain control virus reaction tubes and the water control reaction tube had no amplification, which was a negative result.

Embodiment 2

[0083] The sensitivity result of embodiment 2RT-LAMP detection method

[0084] The starting concentration of porcine ridgevirus genomic RNA was 9.38×10 1 ng / μL, after 10-fold serial dilution, RT-LAMP and common PCR amplification, the results are as follows figure 2 and image 3 As shown, the result shows that the detection limit of the RT-LAMP method of the present invention is about 9.38×10 -9 ng / μL, while the detection limit of common PCR method is 9.38×10 -8 ng / μL.

Embodiment 3

[0085] Fluorescence visualization detection results of embodiment 3 RT-LAMP detection method

[0086] According to the optimized conditions monitored by the turbidimeter, fluorescent dyes were added, reacted at 63°C for 60 minutes, and observed under ultraviolet light. Figure 4 To observe the results, the left tube is the reaction with porcine ridge virus RNA as the template, which is a positive result, and the right tube is a negative control, which is a negative result. The test results show that the established RT-LAMP method can be used conveniently at the grassroots level. You only need to use the kit with the RT-LAMP primers designed by this method. After adding the sample, use a cheap water bath to keep it at 63°C for 60 minutes, and you can quickly observe As a result, without opening the cap, contamination is avoided.

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 a visual reverse transcription loop-mediated isothermal amplification (RT-LAMP) kit for the porcine kobuvirus and application thereof. The kit comprises an RT-LAMP primer, 2*reaction buffer, effective microorganisms (EM), a fluorescent visual detection reagent, ultrapure water and a porcine kobuvirus ribonucleic acid (RNA) template, wherein the RT-LAMP primer comprises outer primers F3 and B3, inner primers FIP and BIP and loop primers LF and LB. The kit is applied to detecting the pathological tissues of the porcine kobuvirus and virus-containing excrement. Specific detection and sensitivity detection verify that the RT-LAMP kit provided by the invention can monitor reaction in real time and quantitatively detect the copy number of the porcine kobuvirus, quickly and accurately obtain the detection results and bring convenience for simply, conveniently, quickly and reliably detecting the porcine kobuvirus.

Description

technical field [0001] The invention relates to the technical field of microbial detection, in particular to a rapid, visualized and real-time quantitative detection of porcine ridge virus reverse transcription loop-mediated isothermal amplification kit and its application. Background technique [0002] Porcine crest virus (Kobuvirus, Kobu) is a single-stranded positive-sense RNA virus belonging to the family Picornaviridae and belonging to the genus Crestavirus. The diameter of the virus particle is about 30nm, and it is icosahedral and symmetrical, without a capsule. In 1998, a new type of RNA virus was found in the stool samples of patients with gastroenteritis in Aichi Prefecture, Japan. It was named Aichivirus according to the place of discovery. In 1997, through genetic analysis, it was confirmed that Aichivirus was a new type of RNA virus. microRNA virus. Because the virion is rugged and ridge-shaped under the electron microscope, it is named cristaevirus. At presen...

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): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/701C12Q1/6844C12Q2531/119C12Q2563/107
Inventor 张宁卢冰霞刘磊何颖秦毅斌郭旋蓝常力赵武李斌段群棚周英宁梁家幸杨思仪蒋冬福苏乾莲
Owner 广西农垦永新畜牧集团金光有限公司
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