TABLE 2.

Primer/probe sequences and conditions for PCR and pyrosequencing of ENTPD1

	Resequencing primer, 5′–3′
Region	Forward	Reverse	Program
Exon 1	TTACAACCTGGAAAAGGCTTC	CTTAAACTGCAAGGGGACTG	P(6–7)
Exon 2	TAAAGGACATGCTGCTTAGGGC	TTCACCCATGATGAAGAAGTG	P(54)
Exon 3	GCAAACAGCATGAAGAGC	CGATAACATAACAACAGCAGC	P(57)
Exon 4	TTGAACACTGAACAGACC	CTCTTATCCTCTTCCTCC	P(57)
Exon 5	CCCCATCTCTTCATTTATTCC	TCATCCATCCATCCATCC	P(6–7)
Exon 6	GTTTCTTTGCCTTAGGAAATCC	TTCTCAGCCTCCCAAGTAGC	P(57)
Exon 7	TGCCTGTTACACAAATCC	TGAGCTGAAGAACCAACC	P(6–7)
Exon 8	AAGCACAAGGGGAAAAAACC	AAACAGGCACAACAGCAGAGCT	P(6–7)
Exon 9	ACTACTTTTTTCCTGGAGGACC	GCAACTGAAAATGCAAGTAGC	P(54)
Exon 10	AACTCTTCTAACTCCTCCAACC	GATTCTTCTTTCAGCCAGC	P(57)
RNA splicing	AGGAGGAAAACAAAAGCTGC	AGATTCCAGGACCTTTAACCC	P(6–7)
Cloning
Promo	ATTAGGTACCCACCGTGCAAAGTAACAGAG	TAATCTCGAGAGGACAGATTGACTGAGGAG	P(6–7)
MCS	ATCTTCCATGGTGGCTTTA	GTGTTGGTTTTTTGTGTGAA	P(6–7)
Pyrosequencing	Biotin-TGCCCTTAGAGGGTTCTTTTC	GTATGGGAGAGATGTCCTCTTTGA
Sequencing	TCCTCTTTGATGCCAG
EMSA	[BIOTEG]CCTTTCAAAGGATTAACCCTTGTTTTGATTT	AAATCAAAACAAGGGTTAATCCTTTGAAAGG

P(6–7): lid 105°C constant; [95°C for 2:00, 95°C for 0:30, 56.7°C for 1:00, 55°C for 0:30] × 9 cycles; 40 cycles [95°C for 0:30, 48.7°C for 0:30, 68°C for 1:00]; 72°C for 10:00; 4°C forever. P(54): lid 100°C; [94°C for 1:30, 94°C for 0:30, 54°C for 0:30; 68°C for 1:30] × 29 cycles, 72°C for 3:00; 4°C forever; P(57) is same as P(54) except annealing is 57°C. For promoter cloning PGL4.23 primers (promo-), underlined nucleotides correspond to KpnI (forward) and XhoI (reverse) restriction sites. For EMSA oligonucleotides, promoter polymorphism rs3814159 alleles are underlined.