Taqman fast pcr reagent

RNA was isolated from HBEpCs using the RNeasy plus kit (Qiagen, Valencia, CA) according to the manufacturer’s instructions, and 1.0 μg RNA was used to generate cDNA with the High capacity ABI-RT PCR kit (Applied Biosystems). RT-PCR reactions were done using the TaqMan FAST PCR reagent (Applied Biosystems) and the ABI 7500 FAST real-time cycler (Applied Biosystems). TaqMan primers for FADD, TNFRSF1, Caspase-3, Caspase-8, and glyceraldehyde phosphate dehydrogenase (GAPDH) were purchased from Thermo Fisher Scientific (Thermo Fisher Scientific, Waltham, MA). Influenza matrix gene expression was quantified with a standard curve and reported as copy numbers of IAV. Gene expression was normalized to GAPDH and reported as fold change. The change in gene expression was calculated thus: fold change = 2^(ddCt), ddCt is the dCt (H1N1-infected) − dCt (mock-infected), where dCt = Ct (detected gene) − Ct (GAPDH) and Ct is the threshold number.

Real time PCR reactions were carried out using the TaqMan FAST PCR reagent (Applied Biosystems) and the ABI 7500 FAST real-time cycler (Applied Biosystems). TaqMan primers for COX6C, Caspase-9, and glyceraldehyde phosphate dehydrogenase (GAPDH) were purchased from Applied Biosystems. Gene expression was normalized using GAPDH as the housekeeping gene, and is reported as fold change. The change in gene expression was calculated using the formula: fold change=2^−(ddCt), ddCt=dCt (H1N1 infected) - dCt (mock infected), where dCt=Ct (detected gene) - Ct (GAPDH) and Ct is the threshold number).