Multiplex RT-PCR (reverse transcription-polymerase chain reaction) detection primer and reagent for swine fever virus (SFV) and porcine reproductive and respiratory syndrome virus (PRRSV)

A technology of RT-PCR and respiratory syndrome, which is applied in the direction of recombinant DNA technology, microbial measurement/inspection, biochemical equipment and methods, etc., which can solve the problems of time-consuming, laborious and high detection costs

Inactive Publication Date: 2013-11-20
曲哲会
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AI Technical Summary

Problems solved by technology

Although some RT-PCR kits for CSFV and PRRSV have been used clinically, they are all single-item RT-PCR methods, that is, three kits are required for the detection of CSFV, classic PRRSV strains and highly pathogenic strains of PRRSV. 3 times, time-consuming and labor-intensive, and the detection cost is also high

Method used

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  • Multiplex RT-PCR (reverse transcription-polymerase chain reaction) detection primer and reagent for swine fever virus (SFV) and porcine reproductive and respiratory syndrome virus (PRRSV)
  • Multiplex RT-PCR (reverse transcription-polymerase chain reaction) detection primer and reagent for swine fever virus (SFV) and porcine reproductive and respiratory syndrome virus (PRRSV)
  • Multiplex RT-PCR (reverse transcription-polymerase chain reaction) detection primer and reagent for swine fever virus (SFV) and porcine reproductive and respiratory syndrome virus (PRRSV)

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Embodiment

[0022] (1) Design and synthesis of primers for multiplex RT-PCR of the present invention

[0023] DNAMAN software was used to compare the homology of 15 strains of PRRSV and 12 strains of CSFV gene sequences in GenBank (see Table 1), and respectively select the two ends of the nsp2 gene deletion site of PRRSV and the 5'UTR gene of CSFV to be conserved In this region, two pairs of primers C1 / C2 and P1 / P2 were designed by using the biological software primer5.0 (the primer pair C1 / C2 can amplify CSFV strains, and the primer pair P1 / P2 can simultaneously identify and amplify PRRSV and HP -PRRSV), using DNAstar for primer-dimer analysis to avoid the formation of stable primer-dimers between primers, and designed amplified PRRSV, HP-PRRSV and CSFV products of 536bp, 452bp and 294bp, respectively. 2 pairs of primers were synthesized by relevant biological companies according to conventional methods.

[0024]

[0025] Table 1 Gene sequences of reference strains for designing PRRSV ...

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Abstract

The invention relates to a multiplex RT-PCR (reverse transcription-polymerase chain reaction) detection primer and reagent for swine fever virus (SFV) and porcine reproductive and respiratory syndrome virus (PRRSV), which can effectively solve the multiplex RT-PCR detection problem of the SFV and the PRRSV. The technical scheme of the invention is as follows: the invention provides the multiplex RT-PCR detection primer for the SFV and the PRRSV, which comprises primer pairs C1/C2 and P1/P2, wherein the primer pair C1/C2 comprises a primer C1 and a primer C2, the primer pair P1/P2 comprises a primer P1 and a primer P2, and the nucleotide sequences of the primer C1, the primer C2, the primer P1 and the primer P2 are SEQ (sequence) ID (identity) NO: 1, SEQ ID NO: 2, SEQ ID NO: 3 and SEQ ID NO: 4 in a sequence table respectively; and after the primer pairs C1/C2 and P1/P2 are used for performing RT-PCR amplification on RNA (ribonucleic acid) of genomes of CSFV (classical swine fever virus) cytotoxicity, PRRSV cytotoxicity and HP-PRRSV (highly pathogenic-porcine reproductive and respiratory syndrome virus) cytotoxicity, CSFVPCR product 294bpSEQ ID NO: 7, PRRSVPCR product 536bpSEQ ID NO: 5 and HP-PRRSVPCR product 452bpSEQ ID NO: 6 are obtained. The invention further provides an application of the primer pairs C1/C2 and P1/P2 in the multiplex RT-PCR detection and testing of the SFV and the PRRSV.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to multiple RT-PCR detection primers and reagents for classical swine fever virus and porcine reproductive and respiratory syndrome virus. Background technique [0002] Classical swine fever (CSF) is an acute, highly contagious infectious disease caused by classical swine fever virus (CSFV). Plague is the most common disease type. The clinical manifestations and pathological changes of the disease are not typical, and it is easy to cause persistent infection in breeding pigs, cause virus-carrying syndrome in sows, and cause immune tolerance in piglets, which is an important reason for the persistence of swine fever in large-scale pig farms . Porcine reproductive and respiratory syndrome (PRRS), also known as "porcine blue ear disease", is caused by porcine reproductive and respiratory syndrome virus (Porcine reproductive and respiratory syndrome virus, PRRSV), mainly in sows An infec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 曲哲会郭晓秋刘涛唐雪峰张广强
Owner 曲哲会
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