When you’re using the ΔΔCt method for relative quantitation, the efficiency of the target and endogenous control must be close to each other (usually within 10% of each other). If the difference is greater than 10%, you need to be aware that when you evaluate fold changes, the unequal PCR efficiencies will correlate to a decrease in the accuracy of the calculated fold change.
We all know that assay efficiency is important, but the “why” can sometimes elude us. Julie from George Washington University was wondering the same thing when she submitted the following question to Ask TaqMan:
“Why is it important? Does the efficiency for all of my targets have to be the same?”
Let’s turn to Sundiep, our director for qPCR assays, in this episode of Ask TaqMan:
Do you have a qPCR question that we can turn into a video? Just ask TaqMan, or tweet your question using the hashtag #askTaqMan.