Inhibition of real-time RT-PCR quantification due to tissue-specific contaminants.
Real-time reverse transcription-polymerase chain reaction (RT-PCR) is currently considered the most sensitive method to study low abundance gene expression. Since comparison of gene expression levels in various tissues is often the purpose of an experiment, we studied a tissue-linked effect on nucleic acid amplification. Based on the raw data generated by a LightCycler instrument, we propose a descriptive mathematical model of PCR amplification. This model allowed us to study amplification kinetics of four common housekeeping genes in total RNA samples derived from various bovine tissues. We observed that unknown tissue-specific factors can influence amplification kinetics but this affect can be ameliorated, in part, by appropriate primer selection.
Tichopad
Ales
A
Institute of Physiology, FML-Weihenstephan, Center of Life and Food Science, Technical University of Munich, Germany.
Didier
Andrea
A
Pfaffl
Michael W
MW
D000818:Animals; D002417:Cattle; D002531:Cerebellum; D003951:Diagnostic Errors; D004340:Drug Contamination; D007700:Kinetics; D008099:Liver; D008962:Models, Theoretical; D018482:Muscle, Skeletal; D009928:Organ Specificity; D012333:RNA, Messenger / Q000032:analysis; D012337:RNA, Ribosomal, 18S / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards*
D012333:RNA, Messenger; D012337:RNA, Ribosomal, 18S