RT-PCR was used to amplify canine lung surfactant protein A (canine lung surfactant protein A, cSP-A) gene, which was ligated into pMD18-T vector. DNAMAN software was used to compare cSP-A gene sequences from different species after sequencing correctly. cSP-A gene was digested by double enzymes and then cloned to pFastbac1 vector to prepare pFastbac1-cSP-A, which was further transformed into DH10Bac competent cells to obtain recombinant bacmid plasmid, and transfected into sf9 cells to generate P1 recombinant baculovirus rBv-cSP-A subsequently; P1 viral stock was propogated serially for three times. The recombinant protein was expressed in sf9 cells infected with P4 rBv-cSP-A, and the expressed product was identified by western Blot and indirect immunofluorescence test. The result showed that the length of cSP-A gene was about 699 bp. The homology of SP-A carbohydrate binding domain among different species was 49.1%-95.5%. Recombinant cSP-A protein was expressed in cells but not secreted out of cells, and its molecular weight was approximately 31.84 kDa by Western Blot and IFA.