Figure 3: Quantitative estimation of vRNA and mRNA levels in H7N9 virus-infected A549 cells.
A549 cells were infected with H7N9 viruses carrying different PB2 adaptation markers, as indicated, at an MOI of 1. Uni-12, Uni-13 and oligo dT primers were used for reverse transcription of vRNA, cRNA and mRNA, respectively. Levels of M1 (a) and NP (b) genes were estimated by quantitative RT-PCR and normalized with the β-actin gene. (c) Verification of specificity of primers used for viral mRNA and vRNA using oligo dT, or uni-12 or uni-13 primed cDNA from NP plasmid transfection and viral particles, respectively, as described in the Methods. The values displayed represent the mean copy number per 105 β-actin copy number±s.d. from three separate experiments. Statistical significance was calculated by the t-test: ***P<0.001 and *P<0.05. h.p.i.: hours post infection.
