Table 2.

Sequences of primers for methylation‐specific polymerase chain reaction and bisulfite genomic sequence

Primer	Primer (forward) (5′→3′)	Primer (reverse) (5′→3′)	Product size and Tm	GenBank accession number
MSP
p16INK4A M‐MSP	TTATTAGAGGGTGGGGCGGATCGC	GACCCCGAACCGCGACCGTAA	150 bp 65°C	X 94154
p16INK4A U‐MSP	TTATTAGAGGGTGGGGTGGATTGT	CAACCCCAAACCACAACCATAA	151 bp 60°C
p21WAF1 M‐MSP	TGTAGTACGCGAGGTTTCG	TCAACTAACGCAACTCAACG	202 bp 53°C	NT_007592
p21WAF1 U‐MSP	TTTTTGTAGTATGTGAGGTTTTGG	AACACAACTCAACACAACCCTA	200 bp 56°C
p27KIP1 M‐MSP	AAGAGGCGAGTTAGCGT	AAA ACGCCGCCGAACGA	177 bp 65°C	S 76986
p27KIP1 U‐MSP	ATGGAAGAGGTGAGTTAGT	AAAACCCCA ATTAAAACA	177 bp 65°C
Bisulfite genomic sequencing
p16INK4A (first PCR)	ATTTTAGGGGTGTTATAT	CTACCTAATTCCAATTCCCCTACA	50°C
p16INK4A (second PCR)	TTTTTAGAGGATTTGAGGGATAGG	TTCCAATTCCCCTACAAA	384 bp, 50°C
p21WAF1 (first PCR)	AATAGTGTTGTGTTTTTTTGGAGAGT	CAAAAATTCCTATACTTATAATCCC	57°C
p21WAF1 (second PCR)	GGGAGGAGGGAAGTGTTTTT	ACAACTACTCACACCTCAACT	251 bp, 52°C

Tm, annealing temperature; M‐MSP, methylation‐specific polymerase chain reaction for the methylated allele; U‐MSP, MSP for the unmethylated allele.