Direct comparison of GAPDH, beta-actin, cyclophilin, and 28S rRNA as internal standards for quantifying RNA levels under hypoxia.

摘要: The appropriate choice of an internal standard is critical for quantitative RNA analyses. As housekeeping genes, GAPDH, beta-actin, cyclophilin, and 28S rRNA are commonly employed as standards with the assumption that their expression levels remain relatively constant in different experimental conditions. We tested this under hypoxia (1% O2, 24 hours) compared to normoxia (20% these 4 genes head using ribonuclease protection assays. Four biologically diverse cell lines respect clonal origin, neoplastic transformation, growth rates were used comparison. Expression constant, independent O2 tension, but cyclophilin varied widely hypoxia. In particular, GAPDH mRNA was increased by 21.2-75.1% hypoxic Increased transcription correlated cancer upregulation factor Hypoxia Inducible Factor-1alpha protein identical These results suggest a reliable control comparative analyses hypoxia; appears particularly unfavorable purpose either or other conditions upregulate HIF-1alpha.