Allele-specific restriction analyses of 8 SNPs
SNPs and primers . | Primer sequence (5′-3′) . | Annealing temperature, °C . | DMSO, % . | PCR product, bp . | Enzyme . | Gel analysis . | Fragment sizes, bp . |
|---|---|---|---|---|---|---|---|
| rs1056286 | 65 | 7.5 | 161 | TaqαI | 2% agarose gel | T = 161 | |
| C = 105 + 56 | |||||||
| F | GTC TGC TGC GGG CGC TGG AC | ||||||
| R | ACT GCG GTG GAG CTC TGC TC | ||||||
| rs722509 | 57 | — | 151 | NsiI | 3% Nusieve | A = 151 | |
| 3:1 agarose gel | T = 129 + 22 | ||||||
| F | GGA ACC CTG ATT CTT TTA GA | ||||||
| R | AAC AAG GTG AGG GGA AAT AC | ||||||
| rs779805 (19A>G)10 | 63 | — | 276 | HaeIII | 4% Nusieve | A = 186 + 90 | |
| 3:1 agarose gel | G = 186 + 45 + 45 | ||||||
| F | GAA ATA CAG TAA CGA GTT GGC CTA GC | ||||||
| R | ACC TCG GCC TCG TCC CAG T | ||||||
| rs779808 | 67 | — | 286 | BstNI | 3% Nusieve | T = 199 + 68 + 19 | |
| 3:1 agarose gel | C = 152 + 68 + 47 + 19 | ||||||
| F | CAT TGC AAC ATC TGC CAC CT | ||||||
| R | GCG ACA GAT CCA GAC TCC AA | ||||||
| rs1678607 | 60 | — | 208 | HpyCH4III | 3% Nusieve | A = 208 | |
| 3:1 agarose gel | C = 147 + 61 | ||||||
| F | CGT ACA AAT ACA TCA CTT CC | ||||||
| R | TGA CGG ACA GCC TAT TTT TG | ||||||
| 1149A>G11 | 50 | — | 110 | AccI | 3% Nusieve | A = 110 | |
| 3:1 agarose gel | G = 89 + 21 | ||||||
| F | CTG CCC ATT AGA GAA GTA TTT | ||||||
| R | AAT TCC CAC TGA ATT ACG TAT A | ||||||
| rs696356 | 58 | 10 | 201 | AvaII | 2% agarose gel | A = 109 + 80 + 12 | |
| C = 189 + 12 | |||||||
| F | CAG TTT CTC TCC AAC AAG GG | ||||||
| R | CCA GGC TGG TCC AGA ACT CC | ||||||
| rs378630 | 67 | — | 225 | BanI | 2% agarose gel | A = 225 | |
| C = 155 + 70 | |||||||
| F | TTA CAG CTC TCC ACT CAC TG | ||||||
| R | GGA GCT CTC TGC AGG GTG AG |
SNPs and primers . | Primer sequence (5′-3′) . | Annealing temperature, °C . | DMSO, % . | PCR product, bp . | Enzyme . | Gel analysis . | Fragment sizes, bp . |
|---|---|---|---|---|---|---|---|
| rs1056286 | 65 | 7.5 | 161 | TaqαI | 2% agarose gel | T = 161 | |
| C = 105 + 56 | |||||||
| F | GTC TGC TGC GGG CGC TGG AC | ||||||
| R | ACT GCG GTG GAG CTC TGC TC | ||||||
| rs722509 | 57 | — | 151 | NsiI | 3% Nusieve | A = 151 | |
| 3:1 agarose gel | T = 129 + 22 | ||||||
| F | GGA ACC CTG ATT CTT TTA GA | ||||||
| R | AAC AAG GTG AGG GGA AAT AC | ||||||
| rs779805 (19A>G)10 | 63 | — | 276 | HaeIII | 4% Nusieve | A = 186 + 90 | |
| 3:1 agarose gel | G = 186 + 45 + 45 | ||||||
| F | GAA ATA CAG TAA CGA GTT GGC CTA GC | ||||||
| R | ACC TCG GCC TCG TCC CAG T | ||||||
| rs779808 | 67 | — | 286 | BstNI | 3% Nusieve | T = 199 + 68 + 19 | |
| 3:1 agarose gel | C = 152 + 68 + 47 + 19 | ||||||
| F | CAT TGC AAC ATC TGC CAC CT | ||||||
| R | GCG ACA GAT CCA GAC TCC AA | ||||||
| rs1678607 | 60 | — | 208 | HpyCH4III | 3% Nusieve | A = 208 | |
| 3:1 agarose gel | C = 147 + 61 | ||||||
| F | CGT ACA AAT ACA TCA CTT CC | ||||||
| R | TGA CGG ACA GCC TAT TTT TG | ||||||
| 1149A>G11 | 50 | — | 110 | AccI | 3% Nusieve | A = 110 | |
| 3:1 agarose gel | G = 89 + 21 | ||||||
| F | CTG CCC ATT AGA GAA GTA TTT | ||||||
| R | AAT TCC CAC TGA ATT ACG TAT A | ||||||
| rs696356 | 58 | 10 | 201 | AvaII | 2% agarose gel | A = 109 + 80 + 12 | |
| C = 189 + 12 | |||||||
| F | CAG TTT CTC TCC AAC AAG GG | ||||||
| R | CCA GGC TGG TCC AGA ACT CC | ||||||
| rs378630 | 67 | — | 225 | BanI | 2% agarose gel | A = 225 | |
| C = 155 + 70 | |||||||
| F | TTA CAG CTC TCC ACT CAC TG | ||||||
| R | GGA GCT CTC TGC AGG GTG AG |
Nusieve 3:1 agarose gel was made by Cambrex BioScience Rockland (Rockland, ME). DMSO indicates dimethyl sulfoxide; F, forward primer; and R, reverse primer.