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 |