Animal insect-borne disease multi-RT-PCR distinguishing and detecting reagent as well as preparation method and application

Abstract

The invention relates to the technical field of biology, in particular to a reagent which can simultaneously distinguish and detect four animal insect-borne diseases as well as a preparation method and application of the reagent. The animal insect-borne disease multi-RT-PCR distinguishing and detecting reagent comprises four pairs of specific primers, and the respective amplification target fragment lengths of Bluetongue virus (BTV), epizootic haemorrhagic disease virus of deer (EHDV), Vesicular stomatitis virus (VSV) and Akabane virus (AKV) are 351bp, 536bp, 300bp and 250bp. The VP7 of the BTV, the EHDV, the VSV and the AKV and a conservative fragment of an N gene are respectively selected as targets, and the primer Express software and the primer prere 5.0 software are applied to deign and combine a primer. An optimal matching and screening test and a multi-RT-PCR test are carried out on a plurality of pairs of designed primers, and four pairs of primers which can carry out the distinguishing and detection on the four animal insect-borne diseases and have high amplification efficiency and good specificity can be obtained by a plurality of reaction condition optimization and comparison tests and verification tests. A kit formed by the reagent can obtain a qualitative distinguishing and detecting result in six hours after a sample is received. The multi-RT-PCR distinguishing and detecting reagent is a sensitive and reliable method for detecting the BTV, the EHDV, the VSV and the AKV in the clinical sample.

Description

technical field [0001] The present invention relates to the field of biotechnology. In particular, a reagent capable of simultaneously identifying and detecting four animal vector-borne diseases, as well as a preparation method and application of the reagent. Background technique [0002] Bluetongue disease (BLU) is a severe animal infectious disease caused by bluetongue virus (BTV) of the reoviridae family (reoviridae) orbivirus genus (Obivirusgenus), and is transmitted by Culicoides , mainly endangers sheep, goats, cattle and other livestock, and is widely prevalent in the world. The World Organization for Animal Health (OIE) lists BLU as an important zoonotic disease. In many national animal trade agreements, BLU is one of the infectious diseases that must be quarantined. In recent years, with climate warming, the incidence of BLU has increased in many European countries, seriously affecting international animal trade. There are 24 serotypes of BTV, and the BTV genome ...

AI Technical Summary

Problems solved by technology

The conventional diagnostic methods for these four diseases are pathogen isolation and identification and serological methods, but these methods have limitations such as low sensitivity, poor specificity, and time-consuming and labor-intensive operations.

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