Total RNA was extracted using the PureLink TM Viral RNA/ DNA Kit (Invitrogen, Carlsbad, USA) according to the manufacturer's instructions, and resuspended in 50 mL of RNase-free water. Reverse transcription-polymerase chain reaction (RT-PCR) was carried out with SuperScript III One-Step RT-PCR System with Platinum Taq DNA Polymerase kit (Invitrogen, Carlsbad, USA) according to the manufacturer's protocols. In brief, a 25-mL reaction mix containing 0.5 mM of each forward and reverse primers and 10 mL of extracted RNA template were used for the RT-PCR. Sets of primers was designed to amplify the complete influenza A(H3N2) HA and NA genes ( Table 2) .