Expression of GAPDH, ?-actin and ?-2-microglobulin genes under chemically induced hypoxic conditions in Hep3b and PC3 cells

Abstract

The appropriate choice of an internal standard is critical for quantitative RNA analyses. As housekeeping genes, GAPDH, ?-Actin and ?-2-Microglobulin are commonly employed as RNA internal standards with the assumption that their expression levels remain relatively constant in different experimental conditions. We have compared the gene expression levels of this genes under normal oxygen level and chemically induced hypoxic conditions using CoCl2 in Hepotoma cell line (Hep3B) and Prostate cell line (PC3). For hypoxic conditions, Hepatoma cells (Hep3B) and prostat cells (PC3) were treated with 150 ?M final consantration of CoCl2 for different time intervals, namely 24, 48 and 72 hours. We used semi-quantitative RT-PCR technique to investigate mRNA level of GAPDH, ?-Actin and ?-2 Microglobulin in Hep3B and PC-3 cell line under chemically induced hypoxic conditions. We also analyzed ?-2 Microglobulin level in HepB cell line by quantitative real-time PCR. As a result we found that the expresion levels of GAPDH and ?-Actin varied widely with hypoxia but expression level of ?-2-Microgloblin RNA were constant in different cell line. We concluded that hypoxic conditions can be optimized in different working conditions and in different cell type.