Detection Of Newcastle Disease Virus Using A SYBR Green I Real Time Polymerase Chain Reaction

Abstract

Summary – A two-step SYBR Green I real time polymerase chain reaction (PCR, real time PCR) for the detection of Newcastle disease virus (NDV) was developed. A melting curve analysis was performed to distinguish specific from non-specific products and primer dimmers. Regardless of different virus pathotypes the melting temperature (Tm) ranged from 86oC to 87oC. The sensitivity of the real time PCR was compared with the reverse transcription (RT) – nested PCR enzyme – linked immunosorbent assay (ELISA, RT-nested PCR ELISA). Whereas the detection limit of the real time PCR was 10 pg DNA, the RT-nested PCR ELISA and conventional PCR could only detect up to 1 ng and DNA, respectively. Thus the real time PCR offers a sensitive, rapid and convenient method for screening large number of NDV specimens.