Rock bream iridovirus (RBIV) is a causative agent of epizootics among cultured rock bream (Oplegnathus fasciatus) in Korea. The structure of the isolated RBIV was observed by an electron microscope, and the virus particles were icosahedral and 120-130 nm in diameter. From the complete genomic DNA sequence of RBIV, the protein encoded in ORF 049L (RBIV-049L) was selected and the property of protein was evaluated with the transmembrane sequence TMHMM 2.0 tool. The ORF 049L gene of RBIV (RBIV-049L) was cloned into pGEX-4T-1 expression vector. The recombinant RBIV-049L was overexpressed in Escherichia coli BL21 (DE3) as a fusion protein (GST-049L, 42 kDa) with a glutathione S-transferase. Antiserum against this recombinant GST-049L protein was prepared in mouse. Dot blot analysis was carried out to identify the reaction abilities and sensitivity of anti-RBIV-049L polyclonal antibody to RBIV-infected rock bream with enzyme linked immunosorbent assay (ELISA) and one-step PCR. These novel RBIV-049L protein and anti-RBIV-049L polyclonal antibody will facilitate the development of more specific and standardized diagnostic techniques. (c) 2006 Elsevier Ltd. All rights reserved.