High Level Expression and Activity Assay of Canine Interferon a Gene
Abstract: With the reverse transcription chain reaction (RT-PCR), the gene of Canine′s interfe ron-alpha (CaIFN-α) was amplified and cloned , from the total RNA in the lymphocytes stimulated with concanavalin A from the peripheral blood of dog. The amplified fragment was inserted into the prokaryotic expressing vector pBV220 and then sequenced. Sequencing result showed that the homology was 100% between the gene we cloned and the one published in GenBank. SDS-PAGE assay showed that the target protein, with the molecular weight of 19kDa, could be expressed in a high level and up to 27.6% of the total protein in E.coli cell, in the form of inclusion bodies. The target protein was added to MDCK cells,which were subsequently infected by the 100 TCID_(50) VSV. The result showed good cytokine activity. The recombinant CaIFN-α activity was up to 5.11×10~6U/㎎.