Objective To construct small interfering RNA（siRNA） eukaryotic expression vector specific for human hnRNP K gene，and to observe its silencing effects on hnRNP K gene in A549 cells.Methods The expression vectors of pSUPER/hnRNP K siRNAa，pSUPER/hnRNP K siRNAc and pSUPER/siRNAn were constructed by gene recombination and then transfected into the A549 lung carcinoma cell line by using Lipofectamine2000（a and c respectively represented A and C fragments in hnRNP K coding sequence contained 19 nts，n represented nonsense fragment as control）.The mRNA and protein were harvested after 24 h and analyzed for the expression of hnRNP K by RT-PCR and Western blotting respectively.Results The siRNA vector targeted to hnRNP K successfully decreased hnRNP K mRNA and protein levels 24 h after transfection in A549 cells.Relative expressed doses of hnRNP K mRNA in lung cancer cells transfected by hnRNP K siRNAa and hnRNP K siRNAc respectively were 0.24±0.53 and 0.28±0.57 after 24 h，which were significantly lower than that in the control group（both Plt;0.01）.The gray scale values of hnRNP K protein were 0.23±0.11 and 0.28±0.09 respectively，which were also significantly lower than those in the control group（both Plt;0.05）.And pSUPER/hnRNP K siRNAa was the most effective one.Conclusion Eukaryotic expression vector of siRNA specific for hnRNP K is successfully constructed，which lays the basis for the function study of hnRNP K gene and its application in the treatment of lung carcinoma.