Identifying Human Host Proteins That Interact With Influenza A Polymerase Proteins Using The Yeast Two-Hybrid System
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Influenza viruses are enveloped, single-stranded RNA viruses which cause 250,000 to 500,000 deaths per year. Understanding how influenza A viral proteins interact with human proteins is essential for identifying factors for species specificity and for developing effective antiviral treatments. The influenza A genome encodes up to 12 proteins, including the polymerase complex consisting of PB1, PB2, and PA. Influenza genes used in this study were amplified from influenza A viruses isolated from swine: H1N1 (PB1-NC, PB2-NC, PA-NC, all of swine origin) and H3N2 (PB1-MN (human origin), PB2-MN (avian origin)). PCR, with a high fidelity polymerase and gene-specific primers, was used for amplification of each of the influenza A genes. PCR products were isolated and used for recombination cloning with the pGBKT7 vector in E. coli cells. Thus far the “bait” pGBKT7 plasmid has been successfully constructed. Yeast 2-hybrid technology (Y2H) will be used to identify which human “prey” proteins interact with the influenza A polymerase proteins.
Heidi Sleister, Mentor