Generally, PLC hydrolyzes PIP2 to generate inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 binds to the IP3-receptor in the membrane of endoplasmic reticulum (ER) and induces the release of Ca2+ into the cytoplasm, whereas DAG, along with Ca2+, activates protein kinase C (PKC). Thirteen PLC isozymes have thus far been cloned from mammalian species, and have been classified into six classes based on their structures and activation mechanisms; PLCδ (delta) (1, 3 & 4), β (beta)(1–4), γ (gamma) (1, 2), ε (epsilon), ζ (zeta), and the recently-discovered η (hepta) (1, 2). These PLC β s are regarded as the sole isoenzymes activated by chemoattractants in the leukocytes. Studies conducted with mice lacking PLCβ2 and PLCβ3 showed that these PLC pathways are critical to chemoattractant-mediated signal transduction and the regulation of protein kinases. Recently, the results of other studies have shown that bradykinin increases IL-6 production in synovial fibroblasts via the bradykinin β2 receptor, PLCβ3, PKCδ, and NF-κB signaling pathways. We cloned a partial fragment of the PLCB3 (878 bp) gene from the brain cDNA of the olive flounder (Paralichthys olivaceus) by designing degenerative primers based on highly conserved domains of PLCB genes, after multiple alignments using ClustalW with full-length cDNAs from previously reported mammalian. In this study, we describe the molecular cloning and sequencing analysis of the olive flounder PLCB3 gene (PoPLCB3), as well as the up-regulation of expression of this gene after lipopolysaccharide (LPS), poly I:C, scuticocilliate and bacterial infection, respectively.The cDNA for olive flounder PLCB3 (PoPLCB3) encodes for a polypeptide of 1,246 amino acids in length containing a well-conserved PH domain, catalytic X and Y domains, a C2 domain. From the sequence information of the BAC library, we assembled a contig containing the whole flounder PLCB3 cDNA sequences, and determined the exon/intron structure of the gene spanning >100,000 bp DNA. Phylogenic analysis and sequence comparison of PoPLCB3 with other PLC isozymes showed a close relationship with the PLCB3 isozyme. Tissue-specific mRNA of PoPLCB3 was expressed predominantly in the kidney, heart and brain tissues. PoPLCB3 gene expression was compared with that of the inflammatory cytokines IL-1β and TNF-α in infected spleen and kidney tissues via real-time RT-PCR assays following stimulation with LPS, poly I:C, scuticocilliate and bacterial infection, respectively