Real-time RT-PCR analysis
Total RNA was extracted from LL29 cells using an RNeasy kit (Qiagen, Hilden, Germany) according to the manufacturer’s protocol. Using a PrimeScript™ II 1st strand cDNA Synthesis kit, samples were reverse-transcribed and THUNDERBIRD® SYBR qPCR Mix, Bio-Rad’s CFX96™ Real-time system, and CFX Manager™ software (Hercules, CA) were used for real-time RT-PCR experiments. Electrophoretic analysis of reaction products was done to confirm specificity. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNA was used as an internal standard. Primers were designed using either Primer3 or Primer-BLAST. Primer sequences will be provided upon request.