Objective:To construct Gadd45γ and its shRNA expression vectors and assess their function on rat glomerular mesangial cell(GMC). Methods:The cds area of Gadd45γ and four 19~21 bp reverse repeated motifs targeting of Gadd45γ gene were synthesized and cloned into eukaryotic expression plasmid pcDNA3.1/HA and pGCsi.U6.neo.GFP. After being screened and confirmed,the recombinant plasmids were transfected into GMCs,then the levels of Gadd45γ protein in rat GMCs were measured using Western blot and immunocytochemistry to prove its expression and find out the optimal shRNA. Results:It was verified by partial nucleotide sequencing and restriction endonuclease digestion that the constructed eukaryotic vectors were correct. The results by Western blot and immunocytochemistry showed that the constructed pcDNA3.1/Gadd45γ plasmid could express correctly,and the optimal shRNA which could effectively silence the target gene,was Gadd45γ shRNA-3. Conclusion:The pcDNA3.1/ Gadd45γ plasmid and its shRNA were constructed successfully. These data provide the foundation for studying biological functions of Gadd45γ gene in the future.