Quantification of Fowl Adenovirus in Tissues

Total DNA was extracted from the liver, spleen, and kidney using an AxyPrep Body Fluid Viral DNA/RNA Miniprep Kit (Axygen, Union City, CA, USA) according to the manufacturer’s instructions. The fragment in the L1 region of the hexon gene of FAdV-4 was used as the target gene, and the chicken ovotransferrin gene was used as a reference gene. Real-time PCR amplification in a 20 μL reaction mixture was performed with Premix Ex Taq (probe qPCR) (TaKaRa, Shiga, Japan) and a QuantStudio 5 system (Applied Biosystems, Foster City, CA, USA). As described above, the final viral load concentration was presented as the copy number per 10⁶ cells [20 (link),21 (link)].