Quantification of multiple human cytochrome P450 mRNA molecules using competitive reverse transcriptase-PCR

DNA and Cell Biology
M R AndersenC J Omiecinski

Abstract

We developed a quantitative competitive reverse transcriptase-polymerase chain reaction (QC RT-PCR) assay to measure mRNA levels of seven human cytochrome P450 (P450, CYP) genes and microsomal epoxide hydrolase (EH) simultaneously. This assay employs an exogenous recombinant RNA (rcRNA) molecule as an internal standard that shares PCR primer and hybridization probe sequences with CYP1A1, CYP1A2, CYP2A6/7, CYP2D6, CYP2E1, CYP2F1, CYP3A4/5/7, and EH mRNA. Because each rcRNA molecule contains several primer sequences, an entire battery of genes that exhibit differential responsiveness to various classes of xenobiotics may be measured simultaneously from one population of cDNA molecules. In this study, we demonstrated the precision and power of the assay using small amounts of human liver total RNA. We also report for the first time quantitative profiles of P450 and EH mRNA abundance in eight human livers. Cytochrome P450 2E1 mRNA maintained the highest abundance (average 6.67 x 10(7) molecules/microg of total RNA) and least variation (13 fold) in all livers examined. Cytochrome P450 1A2, CYP2A6/7, CYP2D6, CYP3A4/5, and EH mRNAs were approximately one order of magnitude less abundant than CYP2E1 transcripts, with CYP2D6 levels exhi...Continue Reading

References

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Citations

Nov 26, 2002·British Journal of Clinical Pharmacology·Anna NordmarkAnders Rane
Feb 25, 2005·Genetic Testing·Lisa A McConnachieRodney J Y Ho
May 30, 2003·Yakugaku zasshi : Journal of the Pharmaceutical Society of Japan·Masuhiro NishimuraTetsuo Satoh
Mar 8, 2013·Seminars in Nephrology·Cortney Bosworth, Ian H de Boer
Aug 5, 2003·Journal of Biochemical and Molecular Toxicology·Stefan Wilkening, Augustinus Bader
Oct 27, 2004·Chemico-biological Interactions·Hollie I Swanson
Feb 7, 2006·Dental Materials : Official Publication of the Academy of Dental Materials·Mario SeissFranz X Reichl
Jul 9, 2010·Drug Metabolism and Pharmacokinetics·Yasuharu NakanishiYasuhiro Uno
Mar 12, 2008·The British Journal of Nutrition·Sinead M BoylanAlastair W M Hay
Dec 11, 2003·Brazilian Journal of Medical and Biological Research = Revista Brasileira De Pesquisas Médicas E Biológicas·L E AlvaresL L Coutinho
Nov 18, 2004·The Journal of Clinical Endocrinology and Metabolism·Ram P GuptaNorman H Bell
Oct 29, 2004·The Journal of Pharmacology and Experimental Therapeutics·R Michael BaldwinAlan R Buckpitt
Jul 18, 2003·Drug Metabolism and Disposition : the Biological Fate of Chemicals·Stefan WilkeningAugustinus Bader
Sep 15, 2001·Archives of Biochemistry and Biophysics·C Rodríguez-AntonaJ V Castell

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