Table 3 The primer pairs, PCR product sizes, restriction enzymes used and the digestion product size in detection of gene polymorphism on pvdhfr and pvdhps gene
Gene/primer pair | PCR product size (bp) | To detectmutation at codon | Restriction enzyme used | Digestion product size (bp) | |
|---|---|---|---|---|---|
Wild type | Mutation | ||||
DHFR Forward: VDF 13NF Reserve: VDF NR58 | 232 | Ile13Leu Pro33Leu Ser58Arg Thr61Met | HaeII SacII AluI Tsp451 | Ile: 232 Pro: 138 and 94 Ser: 167, 40 and 25 Thr: 200 and 32 | Leu: 200 and 32 Leu: 232 Arg: 207 and 25 Met: 232 |
DHFR/ Forward: VDNF57 Reserve: VDF NR | 472 | Phe57Ile/Leu | BsrGI PvuII BstNI BsrI | Phe: 472 Ser: 258 and 214 Ser: 472 Ser: 472 | Ile: 444 and 28 Leu: 472 Asn/Thr: 472 Thr: 257 and 215 Asn: 472 Asn: 253 and 219 Thr: 472 |
DHFR/ Forward: VDT OF Reverse: VDF NR | 608 | Phe57Ile/Leu Ile173Leu | XmnI StyI | Phe: 442 and 166 Ile: 472 and 136 | Ile/Leu: 608 Leu: 438, 97 and 73 |
DHPS/ Forward: VDHPS NF Reverse: VDHPS NR | 703 | Ala383Gly | MspI | Ala: 703 | Gly: 655 and 48 |
DHPS/ Forward: VDHPS553OF Reverse: VDHPSNR | 170 | Ala553Gly | MscI | Ala: 143 and 27 | Gly: 170 |