Real Time PCR Quantification of p21 Upregulation in UV‐Irradiated HaCat Cells

Abstract

The p21 protein, which is upregulated in response to UV‐light, halts cell cycle and can promote apoptosis. Levels of p21 mRNA can be quantified by Real‐Time PCR. We have designed a protocol using Real‐Time PCR to detect the upregulation of p21 transcript in HaCat cells after UV‐irradiation to be used in an undergraduate laboratory course. Cultured HaCat cells were exposed to 15mJ/cm2 UV‐B light, total RNA was extracted and reverse transcribed, and Real‐Time PCR with SYBR Green was performed. Following this level of exposure, levels of p21 were significantly higher in irradiated cells than non‐irradiated cells. This protocol can easily be adapted to be used in any Molecular Biology or Biochemistry Laboratory Course.