Real-time quantitative PCR analysis of gene expression in human colon cancer cells treated with LPS

Relative mRNA levels of 55 genes were evaluated with the SYBR Green qPCR assays after LPS treatment for 8 h. Overall design: Human colon cancer cells (COLO 225) were purchased from ATCC. The cells in triplicate were treated with various concentrations of LPS for 8 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. BCL2 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.

本实验通过SYBR Green qPCR法对LPS处理后8小时的55个基因的相对mRNA水平进行了评估。总体设计为：从ATCC购买了人结肠癌细胞（COLO 225）。将细胞以三重复方式处理以不同浓度的LPS，持续8小时。RNA提取和cDNA合成按先前描述的方法进行。利用Bio-Rad试剂，通过SYBR Green qPCR法对相对mRNA水平进行评估。在本细胞类型中，BCL2 mRNA被选为qPCR分析的内部参照。1% DMSO处理作为样本对照。采用2-ΔCT和2-ΔΔCT相对定量方法，以确定表达量的倍数变化。