Dataset Information

Genetic differentiation of the nucleocapsid protein of Korean isolates of porcine epidemic diarrhoea virus by RT-PCR based restriction fragment length polymorphism analysis.

ABSTRACT: A reverse transcriptase polymerase chain reaction (RT-PCR) based restriction fragment length polymorphism (RFLP) analysis based on the nucleocapsid (N) gene was developed to differentiate between field isolates of porcine epidemic diarrhoea virus (PEDV) and a vaccine strain, J-vac. Thirteen field isolates of PEDV from Korea were distinguishable from the vaccine strain and the prototype PEDV strain CV777 by RFLP using Tru9I. RFLP patterns in 11 of 13 field PEDV isolates were different from the vaccine strain using AspLEI, HgaI and MspR9I. Sequence analysis of the PEDV N gene revealed that Korean field PEDV isolates had 93.6% and 89.6% identity with the vaccine virus at nucleotide and amino acid sequence levels, respectively, suggesting progressive point mutations of the PEDV genome in the field. RFLP analysis of the PEDV N gene is a promising tool for distinguishing field strains from the vaccine-derived virus.

SUBMITTER: Lee C

PROVIDER: S-EPMC7110872 | biostudies-literature | 2008 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Genetic differentiation of the nucleocapsid protein of Korean isolates of porcine epidemic diarrhoea virus by RT-PCR based restriction fragment length polymorphism analysis.

Lee Changhee C Park Choi-Kyu CK Lyoo Young S YS Lee Du Sik du S

Veterinary journal (London, England : 1997) 20070828 1

A reverse transcriptase polymerase chain reaction (RT-PCR) based restriction fragment length polymorphism (RFLP) analysis based on the nucleocapsid (N) gene was developed to differentiate between field isolates of porcine epidemic diarrhoea virus (PEDV) and a vaccine strain, J-vac. Thirteen field isolates of PEDV from Korea were distinguishable from the vaccine strain and the prototype PEDV strain CV777 by RFLP using Tru9I. RFLP patterns in 11 of 13 field PEDV isolates were different from the va ...[more]

PMID: 17728163

Similar Datasets

Project description:BackgroundPorcine epidemic diarrhea virus (PEDV) and porcine delta-coronavirus (PDCoV) are economically important pathogens that cause diarrhea in sows and acute death of newborn piglets. Moreover, the emerging PDCoV was reported to infect children. The current situation is that vaccine prevention has not met expectations, and emergency containment strategies following outbreaks cannot prevent the damages and losses already incurred. Therefore, a more sensitive detection method, that is both convenient and enables accurate and effective sequencing, that will provide early warning of PEDV and PDCoV is necessary. This will enable active, effective, and comprehensive prevention and control, which will possibly reduce disease occurrences.ResultsDuplex nested RT-PCR (dnRT-PCR) is an ideal method to achieve early warning and monitoring of PEDV and PDCoV diseases, and to additionally investigate any molecular epidemiological characteristics. In this study, two pairs of primers were designed for each virus based upon the highly conserved N protein sequences of both PEDV and PDCoV strains retrieved from the NCBI Genbank. After optimization of the reaction conditions, the dnRT-PCR assay amplified a 749-bp fragment specific to PEDV and a 344-bp fragment specific to PDCoV. Meanwhile, the specificity and sensitivity of the primers and clinical samples were tested to verify and establish this dnRT-PCR method. The limit of detection (LoD)for both PEDV and PDCoV was 10 copies/µL. The results showed that among 251 samples, 1 sample contained PEDV infection, 19 samples contained a PDCoV infection, and 8 samples were infected with both viruses, following the use of dnRT-PCR. Subsequently, the positive samples were sent for sequencing, and the sequencing results confirmed that they were all positive for the viruses detected using dnRT-PCR, and conventional RT-PCR detection was conducted again after the onset of disease. As these results were consistent with previous results, a detection method for PEDV and PDCoV using dnRT-PCR was successfully established. In conclusion, the dnRT-PCR method established in this study was able to detect both PEDV and PDCoV, concomitantly.ConclusionsThe duplex nested RT-PCR method represents a convenient, reliable, specific, sensitive and anti-interference technique for detecting PEDV and PDCoV, and can additionally be used to simultaneously determine the molecular epidemiological background.

Dataset Information

Genetic differentiation of the nucleocapsid protein of Korean isolates of porcine epidemic diarrhoea virus by RT-PCR based restriction fragment length polymorphism analysis.

Publications

Genetic differentiation of the nucleocapsid protein of Korean isolates of porcine epidemic diarrhoea virus by RT-PCR based restriction fragment length polymorphism analysis.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets