Methylation-Specific PCR (MSP)

The prediction of CpG island status in the promoter regions of S100A8, MyD88, and NCR3 was conducted using MethPrimer (http://www.urogene.org/). Genomic DNA extraction kits (Vazyme, Nanjing, China) were used to isolate genomic DNA, and agarose gel electrophoresis was performed to determine the integrity of the DNA. In the present study, the isolated DNA specimen was bisulfite-converted using the EpiArt™ DNA Methylation Bisulfite Kit (Vazyme, Nanjing, China). Subsequently, we amplified the samples according to the manufacturer’s instructions, and the primer sequences for MSP amplification are shown in Table 4.

The MSP primer sequences of S100A8 and NCR3.