How much of the first-strand cDNA reaction should I load for PCR?
While the volume is dependent on the starting amount of RNA used for the first-strand synthesis and the abundance of the target gene, we'd recommend starting with 10% of the first-strand reaction for your PCR reaction.
With the Oncomine Focus Assay, Chef-Ready Library, can I perform reverse transcription using the SuperScript VILO cDNA Synthesis Kit on the Ion Chef System?
Reverse transcription needs to be performed manually in the IonCode 96-well PCR plate, using the SuperScript VILO cDNA Synthesis Kit. SuperScript reverse transcriptase enzyme is not supplied in the kit and must be purchased separately. If the RNA was prepared from FFPE tissue and not previously heat-treated, we recommend heating at 80 degrees C for 10 min, then cooling to room temperature.