Real-time quantitative PCR analysis of mouse lung epithelial cells 12 (MLE-12)

MLE12 cells were treated with 8-oxoguanine DNA glycosylase 1 (OGG1) inhibitor with or without TNF to evaluate its effect on pro-inflammatory gene expression. We used SA Biosciences Mouse Inflammatory Cytokines & Receptors PCR Array (PAMM-011Z) to quantitate inflammatory gene expression dependent on OGG1 . Triplicate cultures of cells (-/+OGG1 inhibitor, solvent) were TNF exposed for 30 minutes, RNAs were isolated, pooled (n=3), cDNA were synthesized using Superscript® III First Strand Synthesis System (Invitrogen), mixed with equal amounts of 2X iTaq Universal SYBR Green Supermix (Bio-Rad) and 20 μl of reaction mixture was added to each well of the PAMM-011ZA array. The qRT-PCR was performed using an ABI PRISM® 7000 Sequence Detection System as recommended by SA Biosciences.