De-modification of RNA results in higher efficiency during subsequent QPCR. RNA was isolated from FFPE material according to our own protocol and compared to intact RNA derived of FF tissue. RNA samples were reverse transcribed without previous de-modification (labeled "no") or after de-modification at room temperature (1), 94°C and pH 8.0 (2) or 94°C and pH 5.0 (3). Each RNA was tested by QPCR using three amplicons for IGBP5. Primers used code for short (60 bp, □), medium-size (109 bp, ◇) or long amplicons (147 bp, △). Shown are raw Ct values from intact RNAs from FF material and from RNAs derived of FFPE material of the same tumors. The benefit of de-modification is visualized as delta Ct values. They are indicated for short and long amplicons (dotted lines).
Oberli et al. BMC Medical Genomics 2008 1:9 doi:10.1186/1755-8794-1-9