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

RT-LAMP primer set and kit for detecting bluetongue virus, epizootic haemorrhagic disease virus and Palyam virus

A technology of RT-LAMP and epidemic hemorrhagic disease, which is applied in the field of RT-LAMP primer sets and kits, can solve problems such as limitations and inability to realize on-site rapid detection, and achieve short reaction time, great promotion value, high specificity and The effect of sensitivity

Pending Publication Date: 2021-09-03
YUNNAN ANIMAL SCI & VETERINARY INST
View PDF4 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, for BTV, EHDV and PALV, a variety of nucleic acid detection methods including RT-PCR and qRT-PCR have been established, but the above methods require the use of complex and expensive instruments such as PCR machines, so they are mainly limited to laboratory testing and cannot be implemented on-site Quick check

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
  • RT-LAMP primer set and kit for detecting bluetongue virus, epizootic haemorrhagic disease virus and Palyam virus
  • RT-LAMP primer set and kit for detecting bluetongue virus, epizootic haemorrhagic disease virus and Palyam virus
  • RT-LAMP primer set and kit for detecting bluetongue virus, epizootic haemorrhagic disease virus and Palyam virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0143] Embodiment 1, primer design and preparation

[0144] Download the BTV Seg-5 sequence, EHDV Seg-5 sequence and PALV Seg-7 sequence from GenBanK, use MEGA5.0 for comparison and analysis, select highly conserved regions, and use Primer Exporer V5 online software to design 3 sets of RT-LAMP amplification primers.

[0145]Primer set for identifying BTV, including two outer primers (BTV-F3, BTV-B3), two inner primers (BTV-FIP, BTV-BIP) and two loop primers (BTV-LB, BTV-LF) Composition; primer set for identifying EHDV, including two outer primers (EHDV-F3, EHDV-B3), two inner primers (EHDV-FIP, EHDV-BIP) and two loop primers (EHDV-LB, EHDV- LF) composition; primer set for identifying PALV, including two outer primers (PALV-F3, PALV-B3), two inner primers (PALV-FIP, PALV-BIP) and one loop primer (PALV-LF) The primer sequences of each set are shown in Table 1.

[0146] Table 1 BTV, EHDV and PALV group-specific RT-LAMP detection primer information

[0147]

[0148] Primers...

Embodiment 2

[0149] Embodiment 2, detection method establishment

[0150] 1. Samples to be inspected

[0151] Virus to be tested 1: BTV-9 strain and monitored animal blood samples from which the virus was isolated (recorded in the following literature: Li Zhanhong, Wang Jinping, Yang Heng, etc. The first isolation of bluetongue virus serotype 9 in my country[ J]. Journal of Animal Husbandry and Veterinary Medicine, 2019, 50(2):354-363.).

[0152] Blood sample to be tested 1: Sentinel cattle EDTA anticoagulated blood sample from which the BTV-9 strain was isolated (recorded in the following literature: Li Zhanhong, Wang Jinping, Yang Heng, etc. The first bluetongue virus serotype 9 strain in my country Separation [J]. Journal of Animal Husbandry and Veterinary Medicine, 2019, 50(2):354-363.).

[0153] Virus 2 to be tested: EHDV-10 strain (recorded in the following literature: Li Zhanhong, Xiao Lei, Yang Zhenxing, et al. Isolation and identification of bovine epidemic hemorrhagic disease vir...

Embodiment 3

[0175] Embodiment 3, sensitivity

[0176] 1. Preparation of Standards

[0177] Use the T7 in vitro transcription kit to transcribe the ssRNA template of the target gene, use the NanoDrop 2000 nucleic acid concentration analyzer to measure the concentration of in vitro transcribed ssRNA, and calculate the nucleic acid copy number. The ssRNA with the determined copy number was serially diluted 10 times as a standard.

[0178] 2. Sensitivity test

[0179] (1) The samples to be tested are: BTV-Seg-5ssRNA (copy number: 4.5×10 10 copy / μL), EHDV-Seg-5ssRNA (copy number: 6.2×10 10 copy / μL) and CHUV-Seg-7ssRNA (copy number: 2.3×10 11 copy / μL);

[0180] (2) Dilution of samples to be tested: each sample to be tested was diluted 10 times with RNase-free water, and BTV-Seg-5ssRNA was diluted to 4.5×10 0 copies / μL to 4.5 x 10 5 copies / μL, EHDV-Seg-5ssRNA was diluted to 6.2×10 0 copies / μL to 6.2×10 5 copies / μL, PALV-Seg-7ssRNA was diluted to 2.3×10 0 copies / μL to 2.3×10 5 copies / μ...

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
Copy numberaaaaaaaaaa
Login to View More

Abstract

The invention relates to an RT-LAMP primer set and kit for detecting bluetongue virus, epizootic haemorrhagic disease virus and Palyam virus. The RT-LAMP primer set comprises a primer set I, a primer set II and a primer set III, wherein the primer set I consists of BTV-F3, BTV-B3, BTV-FIP, BTV-BIP, BTV-LB and BTV-LF; the primer set II consists of EHDV-F3, EHDV-B3, EHDV-FIP, EHDV-BIP, EHDV-LB and EHDV-LF; and the primer set III consists of PALV-F3, PALV-B3, PALV-FIP, PALV-BIP, and PALV-LF. The RT-LAMP detection method provided by the invention has high specificity and sensitivity, can quickly and accurately detect BTV, EHDV and PALV, and has great popularization value.

Description

technical field [0001] The invention belongs to the technical field of detection of veterinary infectious diseases, in particular to an RT-LAMP primer set and a kit for visual detection of bluetongue virus, zoonotic hemorrhagic disease virus and paliam virus. Background technique [0002] Bluetongue virus (BTV), Epizootichaemorrhagic disease virus (EHDV) and Palyam virus (PALV) are ruminant arboviruses transmitted by Culicoides and belong to respiratory A member of the Orbivirus genus of the Enteroviridae family. The genomes of BTV, EHDV, and PALV are all composed of 10 segments (Seg-1~Seg-10) of double-stranded RNA (Double stranded RNA, dsRNA), with a size of about 20kb, and have close genetic relationship in evolution. [0003] BTV-infected sheep can cause obvious clinical symptoms, manifested as high fever, weight loss, lip congestion, swelling and erosion, nasal cavity and gastrointestinal mucosal bleeding ulcers, and lameness caused by shedding of hooves and crowns; wh...

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): C12Q1/70C12Q1/6844C12N15/11
CPCC12Q1/701C12Q1/6844C12Q2521/107C12Q2531/119
Inventor 李占鸿杨恒廖德芳杨振兴李卓然李华春
Owner YUNNAN ANIMAL SCI & VETERINARY INST
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