Table 1.

Primer and probe nucleotide sequences used in this study.

GENE EXPRESSION qPCR
GENE	PRIMER SEQUENCE	PROBE SEQUENCE
t-PA	Fw 5′-GGC CTT GTC TCC TTT CTA TTC G-3′ Rv 5′-AGC GGC TGG ATG GGT ACA G-3′	5′-TGA CAT GAG CCT CCT TCA GCC GCT-3′
GAPDH	Fw 5′-CCA CAT CGC TCA GAC ACC AT-3′ Rv 5′-CCA GGC GCC CAA TAC G-3′	5′-AAG GTG AAG GTC GGA GTC AAC GGA TTT G-3′

ChiP qPCR
REGION	PRIMER SEQUENCE	LOCATION*
t-PA promoter	Fw 5′-ACC CCC TGC CTG GAA ACT TA-3′ Rv 5′-GGT ACA GAA ACC CGA CCT ACC A-3′	−46 to +92

BISULFITE SEQUENCING
REGION		#	PRIMER SEQUENCE	LOCATION*
t-PA promoter 1	Outer	1	Fw 5′-TTT GAA AAG GTG TTA GTA AG-3′ Rv 5′-ACC ACT AAA AAA ACA AAA CC-3′	−721 to −382
	Inner	2	Fw 5′-TAA GGG AAA TGG TTT GTT TA-3′ Rv 5′-CTACRATAAAAAATACCCCCATA-3′	−705 to −416
t-PA promoter 2	Outer	3	Fw 5′-TTT GGG TTT ATT TAA GGG GAT GT-3′ Rv 5′-AAA AAT TTT CTC TCC AAC CCT AAA C-3′	−577 to +156
	Inner	4	Fw 5′-GAG GTT ATT TAT TGT AGT TTT GTA TTT TAT-3′ Rv 5′-CAA CTC TAA ACT CCC CAC AAC TC-3′	−535 to +118
t-PA promoter 3	Outer	5	Fw 5′-TTA GGA TTT TAA AGG AAG ATG ATT TTT AA-3′ Rv 5′-AAA AAA ACA AAC CCC AAA ATA CAA-3′	−176 to +222
	Inner	6	Fw 5′-AAA GGA AGA TGA TTT TTA AGG TTT TAT TT-3′ Rv 5′-AAA ATT TTC TCT CCA ACC CTA AAC T-3′	−166 to +155

Notes:

^*Location with respect to the transcription start site of the t-PA gene. Top panel: Primers and TaqMan probes used in the real-time RT-PCR. mRNA of the t-PA gene was converted to complementary DNA and quantitatively amplified together with the reference gene GAPDH. Middle panel: Primes sequences used for SYBR Green real-time PCR quantification of chromatin immunoprecipitation (ChIP) samples. Bottom panel: Primer sequences used in the methylation analysis of the t-PA promoter. Three overlapping regions of the t-PA promoter (t-PA promoter 1, 2 and 3) were amplified in a nested PCR. The reaction was run in two stages with outer and an inner primers. Locations are given according to the NCBI Reference Sequences for t-PA. Fw denotes forward, Rv denotes reverse.