Real-time quantitative PCR analysis of gene expression in mouse macrophages treated with bacterial endotoxin lipopolysaccharide

Relative mRNA levels of 27 genes were evaluated with the SYBR Green qPCR assays after LPS treatment for 2-24 h. Overall design: Mouse macrophages (RAW264.7 cells, TIB-71) were established from a tumor in a male mouse induced with the Abelson murine leukemia virus (ATCC). The cells in triplicate were treated with various concentrations of LPS for 2, 8 and 24 h. RNA isolation and cDNA synthesis were performed as described previously. Relative mRNA levels were evaluated with the SYBR Green qPCR assays using Bio-Rad reagents. Rpl32 mRNA was selected as the internal reference for qPCR analysis for this cell type. 1% DMSO treatment was the sample control. The 2-ΔCT and 2-ΔΔCT method of relative quantification was used to determine the fold change in expression.