Quantitative determination of gene expression by competitive reverse transcription-polymerase chain reaction in degraded RNA samples.
Quantitative competitive reverse transcription-polymerase chain reaction has become a powerful tool for determining the amount of mRNA in cultured cells as well as in tissue. To ensure the reliability of the analysis, RNA with high purity and integrity is needed. However, when analyzing RNA samples from tumor biopsies, RNA degradation is often an inevitable problem. This causes differences in sample quality and furthermore adversely affects the quantification of gene expression. Here, we demonstrate that in partially degraded RNA samples different mRNAs are degraded to the same extent. Normalizing the expression level of a specific gene to that of a constitutively expressed gene in the same sample allows the relative quantification of this specific gene. Thus, the comparison of gene expression in RNA samples with varying integrity is possible.
Tong
D
D
Division of Gynecology, University of Vienna, Vienna, Austria.
Schneeberger
C
C
Leodolter
S
S
Zeillinger
R
R
D015870:Gene Expression; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000737:chemistry*; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases