Total RNA of infected Vero E6 cells was extracted with the RNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. Reverse transcription and PCR amplification were performed with an iTaq Universal One-Step RT-qPCR Kit (Bio-Rad Laboratories, Hercules, CA). Real-time PCR was performed in a StepOnePlus Real-Time PCR System (Applied Biosystems, Foster City, CA). The primer pairs used to amplify the viral RNA were as follows: GAPDH-F: 5′-GAAGGTGAAGGTCGGAGTC-3′; GAPDH-R: 5′-GAAGATGGTGATGGGATTTC-3′;53 MERS-CoV-F: 5′-CCACTACTCCCATTTCGTCAG-3′; MERS-CoV-R: 5′-CAGTATGTGTAGTGCGCATATAAGCA-3′.54 The MERS RNA levels were normalized to that of GAPDH and compared between MERS-CoV groups at 24 h.p.i. and at 48 h.p.i.