Table 1.

PCR primers, probes, concentrations, and conditions. Polymorphic site underlined in probe sequences.

Gene and Polymorphism (dbSNP RS no.)	Primers and Probes, 5’–3’ Sequence	Concentration	Conditions
	TaqMan
MTHFR 677C > T (1801133)	F: GCA GGG AGC TTT GAG GCT GAC C	0.5 μM	92°C 30 s
	R: TGG GGC AAG TGA TGC CCA TGT	0.5 μM	56°C 1 min
	*T: 6FAM-ATG AAA TCG <U>A</U>CT CCC GC-MGBNFQ	50 μM	50 cycles
	*C: VIC-ATG AAA TCG <U>G</U>CT CCC GC-MGBNFQ	100 μM
MTHFR 1298A > C (1801131)	F: GAG GAG CTG CTG AAG ATG T	0.5 μM	92°C 30 s
	R: CGA GAG GTA AAG AAC GAA GA	0.5 μM	56°C 1 min
	*C: 6FAM-AGA CAC TT<U>G</U> CTT CAC T-MGBNFQ	50 μM	50 cycles
	*A: VIC-CAA AGA CAC TT<U>T</U> CTT C-MGBNFQ	50 μM
MTR 2756A > G (1805087)	F: AGT GTT CCC AGC TGT TAG ATG A	0.5 μM	92°C 30 s
	*R: TGT TTC TAC CAC TTA CCT TGA GAG ACT	0.5 μM	60°C 1 min
	*G: 6FAM-ACA GG<U>G</U> CCA TTA TG-MGBNFQ	50 μM	50 cycles
	*A: VIC-ATT AGA CAG G<U>A</U>C CAT TAT G-MGBNFQ	100 μM
MTRR 66A > G (1801394)	*F: CAT GCC TTG AAG TGA TGA GG	0.5 μM	92°C 30 s
	*R: GAT CTG CAG AAA ATC CAT GTA CCA	0.5 μM	60°C 1 min
	*G: 6FAM-CTT GCT CAC A<U>C</U>A TTT-MGBNFQ	50 μM	50 cycles
	*A: VIC-TGC TCA CA<U>T</U> ATT TC-MGBNFQ	100 μM
	Size Difference PCR
CBS 844ins68	F: TAT TGG CCA CTC CCA TAA TAG A	0.4 μM	94°C 5 min
	R: CGG CTC TGC GAG GAT GGA CCC TT	0.4 μM	35 cycles of
			94°C 1 min
			55°C 1 min
			72°C 1 min
TYMS 1494de16 (16430)	F: CAT GAT GTA GAG TGT GGT TAT G	0.4 μM	94°C 2 min
	R: GAA TGA ACA AAG CGT GGA	0.4 μM	35 cycles of
			94°C 30 s
			51 °C 30 s
			72°C 30s then
			72°C 5 min
DHFR c.86+60_78	F1: CCA CGG TCG GGG TAC CTG GG	0.4 μM	94°C 4 min
	F2: ACG GTC GGG GTG GCC GAC TC	0.4 μM	35 cycles of
	R: AAA AGG GGA ATC CAG TCG G	0.8 μM	94°C 55s
			62°C 55 s
			72°C 55 s
			then 72°C 12 min

^*Originated from SNP500Cancer website²⁶.