Abstract: Abstract: Objective To establish a simple, rapid and inexpensive method to determine the methylation status of eight key clock genes, BMAL1, BMAL2, CLOCK, NPAS2, PER1, PER2, CRY1 and CRY2. To examine the methylation of clock promoters in five peripheral tissues with the developed method. Methods Genome DNA was deaminated by Sodium metabisulfite solution and hydroquinone. The two sets of PCR primers for unmenthylated and methylated DNA, respectively, were used to amplify the promoter of clock genes. Polymerase chain reaction (PCR) products were then loaded and electrophoresed on 3% agarose gels. The PCR products for each reaction were sequenced directly. Results Specific amplicons with correct size were amplified. PCR products were also confirmed by sequencing. Conclusion In the present study, a new method of detection of the methylation of clock genes was successfully established; it may apply a new technique for clock genes promoter methylation detection. All of the clock promoters are free from methylation in the adult mouse.

Key words: Key words: clock gene, promoter, methylation specific PCR