Construction Of Sirnaeukaryotic Expressionvectorformouse SPK1Gene Andresearchof Transfecting It Into Lewis Lung Cancercells

Objective: To construct a mouse SPK1gene-targeting siRNA expression vector and totransfect it into LLC cells，then provide a technological means to study the relationshipbetween SPK1with non-small cell lung cancer.Methods: To select the specific siRNA sequences of mouse SPK1gene which hadproven by the literature, according to the above-mentioned sequence, we synthesis thestencil sequence, and then connect it with the linearized plasmid vector cut by doubleenzyme (Age I and EcoR I), sequencing. We transfect the plasmid into LLC cells withlipofectamine2000. To observe the cells by fluorescence microscope, detect the efficiencyof transfection by flow cytometry and detect the expression levels of SPK1protein in LLCcells by western blot after transfecting.Results: We have successfully constructed a mouse SPK1gene-targeting siRNAexpression vector, and effectively reduced the SPK1expression of LLC cells aftertransfecting.Conclusion: The siRNA eukaryotic expression vector for mice SPK1gene weconstructed can be stably transfected into LLC cells to inhibit the expression of SPK1protein，and can be useful in further studies the realtionship between SPK1and non-smallcell lung cancer.