With real-time PCR (qPCR), target-gene expression levels are often normalized to the expression of a reference gene to obtain relative quantification of expression. This normalization strategy corrects for differences in the amount of input RNA and variation in reaction efficiency. A housekeeping gene that is stably expressed in all the samples being tested is commonly used as the reference. However, in some experimental systems, it may be difficult to identify a gene with stable expression. In such cases, an external standard RNA can be added to the reaction to serve as a reference.