Heterologous Expression And Enzymatic Activity Identification Of CYP6CW1in Nilaparvata Lugens

The Bac-to-Bac baculovirus expression system is a sort of gene expression system that allows the rapid and efficient generation of recombinant baculovirus DNA by site-specific transposition in Escherichia coli. We previously established co-expression parameters for Brown Planthopper, Nilaparvata lugens Stal (Homoptera:Delphacidae) CYP6CW1and housefly Mcusca dometica NADPH-Cytochrome P450reductase (NCPR) in baculovirus-infected Sf9cells that allow for efficient expression of both components and significantly enhance metabolic turnover of this insect P450’s substrates. Fresh insect Sf9cells were infected with the recombinant virus containing NCPR and CYP6CW1to express the target protein differently and together. Result showed that the recombinant bacmid expression vector, MH, were constructed successfully. What’s more the NCPR gene and CYP6CW1gene were both expressed in Sf9insect cells. The specific bands (about76.4kD and61.8kD) were detected by SDS-PAGE and confirmed by Western blot analysis separately. The co-expression ended, we got the CYP6CW1recombinase with metabolic activity. The enzyme activity was determined using Phenobarbitalum as a substrate which was determinated by High performance liquid chromatography (HPLC). The result revealed that CYP6CW1can metabolize Phenobarbitalum, suggesting that the P450gene may play an important role in detoxification of rice allelochemicals and in strengthening tolerance of brown planthopper to host resistane.