Total RNA was extracted from the brain, lung, spleen, and bursa of Fabricius of H5N6-inoculated chickens and mock-infected chickens at 3 DPI using the Takara MiniBEST Universal RNA Extraction Kit (Takara Bio Inc., Tokyo, Japan) following the manufacturer’s instructions. Total RNA (1 μg) was reverse-transcribed with the PrimeScriptTM II 1st Strand cDNA Synthesis Kit (Takara Bio Inc.) and stored at -20°C for further study.