Cyprinid herpesvirus 2(CyHV2) is a fatal pathogen for some species of the Cyprininae fish and, furthermore, is responsible for negatively impacting the goldfish industry worldwide. In order to develop a polymerase chain reaction (PCR) capable of detecting CyHV2, 2 specific primers and 9 overlapping oligo primers were designed based upon the nucleotide sequence information of CyHV2 published in GenBank (accession no:HM014349), and a 413 bp DNA fragment of the CyHV2 DNA-dependent DNA polymerase gene was synthesized in vitro by using an overlap extension PCR to construct the recombinant plasmid,pMD19-T-CyHV2. Then, a primary PCR method was established after set of serial tests, including reaction conditions optimization test, sensitivity test, and specificity test. The final results indicate that this developed PCR assay is a rapid method that maintains both a strong specificity and a high sensitivity for detecting CyHV2. The PCR detection limit could reach approximately 62 copies of the cloned viral genomic fragments (pMD19-T-CyHV2) as well as resulted in no amplifications for Aeromonas veronii, Aeromonas hydrophila, Pseudomonas fluorescens and Streptococcus, which are common pathogens isolated from fish by this detection approach.
Cyprinid herpesvirus 2, PCR, overlap extension PCR
© The Author(s) 2014. Open Access. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License which permits unrestricted use, sharing, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.