Growth arrest and DNA damage inducible 45-beta (Gadd45B) is essential for mitogen-activated protein kinases (MAPK) activities, and involved in regulating growth, apoptosis, and DNA demethylation. In the present study, the cDNA of gcGadd45Ba and gcGadd45Bb in grass carp was identified. And the expression levels show that they were widely distributed in the tested tissues and showed significant immune responses both in vitro and in vivo after challenge with bacteria and pathogen-associated molecular patterns (PAMPs). Overexpression of Gadd45B significantly induced the expression of pro-inflammatory cytokines (IL-1β, IL-8, and TNF-α) and enhanced the phagocytosis activation of grass carp blood cells. These results indicate that Gadd45B plays an important role in innate immune responses.