Genotyping method for CYP2C8 polymorphisms by PCR-RFLP or AS-PCR
Polymorphisms | Primer Set | PCR-RFLP (Restriction Enzyme) or AS-PCR | Concentration of MgCl2 | Temperature for Annealing | Fragment Length | Electrophoresis |
|---|---|---|---|---|---|---|
| mM | °C | bp | ||||
| A805T (1269F, CYP2C8*2) | 5S | RFLP (MboI) | 2.0 | 48 | wt: 69, 65, 33 | 12% Polyacrylamide |
| 5AS | mt: 98, 69 | |||||
| G416A (R139K, CYP2C8*3) | 3S | RFLP (BseRI) | 2.0 | 55 | wt: 310, 157 | 2% Agarose |
| 3AS | mt: 467 | |||||
| C792G (1264M, CYP2C8*4) | 5S | RFLP (TaqI) | 2.0 | 48 | wt: 83, 53, 31 | 12% Polyacrylamide |
| 5AS | mt: 136, 31 | |||||
| A475 del (Frameshift, CYP2C8*5) | 3S | AS-PCR | 1.5 | 51 | 370 | 2% Agarose |
| A475del-wt or A475del-mt | ||||||
| C389A (T130N) | 3S | AS-PCR | 1.5 | 53 | 288 | 2% Agarose |
| T130N-wt or T130N-mt | ||||||
| G462T (E154D) | 3S | RFLP (EcoNI) | 2.0 | 55 | wt: 388, 129 | 2% Agarose |
| 3AS | mt: 467 | |||||
| T579A (N193K) | N193K-S | AS-PCR | 1.5 | 50 | 382 | 2% Agarose |
| N193K-AS-wt or N193K-AS-mt | ||||||
| A746G (L249R) | K249R-S | AS-PCR | 1.5 | 55 | 510 | 2% Agarose |
| K249R-AS-wt or K249R-AS-mt | ||||||
| T1196C (L390S) | L390S-wt or L390S-mt | AS-PCR | 2.0 | 55 | 254 | 2% Agarose |
| 8AS | ||||||
| C1210G (P404A) | P404A-wt or P404A-mt | AS-PCR | 2.0 | 55 | 215 | 2% Agarose |
| 8AS | ||||||
| A1232T (H411L) | 8S | RFLP (EaeI) | 2.0 | 50 | wt: 429, 178 | 2% Agarose |
|
| 8AS |
|
|
| mt: 607 |
|
bp, base pairs; wt, wild type; mt, mutant type.