qRT-PCR analysis of p53, p21, and MDM2 expression

qRT-PCR was performed as our previous publication [14 (link), 17 (link)]. Total RNA was isolated from the cells treated with HST or DMSO for 48 h using TRIzol (Life Technologies, Carlsbad, CA, USA). cDNA was synthesized from 1 μg of total RNA using StarScript II First-strand cDNA Synthesis Mix (Genstar, Beijing, China). According to the manufacturer’s instructions, qPCR was further conducted with aliquots of cDNA samples mixed with 2 × RealStar Green Fast Mixture (Genstar, Beijing, China) using a CFX96™ Real-Time PCR Detection System (BIO-RAD, Hercules, CA, USA). GAPDH was used as the reference gene for normalization using the 2^−△△Ct method. The sequences of primers were as follows:

p53, Fw 5′-TAACAGTTCCTGCATGGGCGGC-3′,

Re 5′-AGGACAGGCACAAACACGCACA-3′;

p21, Fw 5′-TGTCCGTCAGAACCCATGC-3′,

Re 5′-AAAGTCGAAGTTCCATCGCTC-3′;

MDM2, Fw 5′-AGTAGCAGTGAATCTACAGGGA-3′,

Re 5′-CTGATCCAACCAATCACCTGAAT-3′;

GAPDH, Fw 5′-CATGAGAAGTATGACAACAGCCT-3′,

Re 5′-AGTCCTTCCACGATACCAAAGT-3′;