2.5. Quantitative Real Time Polymerase-Chain Reaction

For the analysis of the mRNA level of target genes of p53 in PGA₂-treated cells, quantitative real time polymerase-chain reaction (qPCR) was performed as follows. First-strand cDNA was synthesized from total RNA using PrimeScript^TM RT reagent Kit (Takara Korea Biomedical Inc., Seoul, Korea). First-strand cDNA was then amplified by specific primers against target genes of p53 using SYBR FAST qPCR Kit (KAPAbiosystems, Woburn, MA, USA) on ABI 7300 Real-Time PCR System (Applied Biosystems, Carlsbad, CA, USA). GAPDH mRNA level normalized each mRNA level of p53-target genes in the same sample and their relative changes among samples were calculated by the ΔΔCt method [22].