Quantifying Ovarian Cancer Gene Expression

Subconfluent ovarian cancer cells were treated with varied concentrations of monesin for 48 h. Total RNA was isolated from the treated cells by using TRIZOL Reagents (Invitrogen) and subjected to reverse transcription reactions with hexamer and M-MuLV reverse transcriptase (New England Biolabs, Ipswich, MA). Such cDNA products were used as PCR templates. The qPCR primers were designed by using Primer3 program69 (link) and used to amplify the genes of interest (Supplemental Table 1). The TqPCR were carried out by using the SYBR Green-based qPCR analysis on a CFX-Connect unit (Bio-Rad Laboratories, Hercules, CA), as described24 (link). The qPCR reactions were done in triplicate. GAPDH was used as a reference gene.