Table 1.
The List of Candidate CYP2D6 and CYP2E1 Genes Selected in the Present Study with Details of PCR and RFLP Procedures Including Details of Primers and Their Sequences, Restriction Enzymes and Expected base Pairs of Expected PCR Products
| Gene/ Genotype |
rs number | Nucleotide change |
Primer Sequence (Forward/Reverse) |
PCR Conditions |
PCR product size |
Enzyme / Digestion conditions |
Dominant Wild type (bp) |
Heterozygous (bp) |
Recessive Mutant (bp) |
|---|---|---|---|---|---|---|---|---|---|
|
CYP2D6*3 2549delA |
rs35742686 | (A>- (del A) | FP: 5’-ATG AGC TGC TAA CTG AGC CC -3’ RP: 5’- CCG AGA GCA TAC TCG GGA C -3’ |
95°C for 5 min , 30 cycles of 95°C- 30 sec, 54°C- 45 sec, 72°C- 60 sec , final extension 72°C for 5 min |
270 bp | 1 Unit of MspI Incubation at 37˚C for 16h |
188 bp, 82 bp | 188 bp, 168 bp 20 bp |
168 bp, 20 bp |
|
CYP2D6*4 1846G>A |
rs3892097 | (G>A) | FP: 5’-GCT TCG CCA ACC ACT CCG-3’ RP: 5’-AAA TCC TGC TCT TCC GAG GC-3’ |
95°C for 5 min, 30 cycles of 95°C- 20 sec, 57°C- 20 sec, 72°C- 20 sec ,final extension 72°C for 5 min |
334 bp | 1 Unit of BstO1 Incubation at 37˚C for 1h |
230 bp, 105 bp | 334 bp, 230 bp 105 bp |
334 bp |
|
CYP2D6*10 100C>T |
rs1065852 | (C>T) | FP: 5’- GTG CTG AGA GTG TCC TGC C -3’ RP: 5’- CAC CCA CCA TCC ATG TTT GC -3’ |
95°C for 5 min, 30 cycles of 95°C- 60 sec, 53°C- 60 sec, 72°C- 60 sec, final extension 72°C for 5 min |
344 bp | 1 Unit of HphI Incubation at 37˚C for 16h |
282 bp, 62 bp | 282 bp, 182 bp, 100bp, 62bp |
182 bp, 100 bp 62 bp |
|
CYP2D6*17 1023C>T |
rs28371706 | (C>T) | FP: 5’- CGG TGG TCG TGC CTC AAT G -3’ RP: 5’- CCC GGG TCC CAC GGA AAT CT -3’ |
95°C for 5 min, 30 cycles of 95°C- 30 sec, 55°C- 45 sec, 72°C- 60 sec, final extension 72°C for 5 min |
167 bp | 1 Unit of HphI Incubation at 37˚C for 1h |
167 bp | 167 bp, 88 bp 55bp |
88 bp, 55 bp |
|
CYP2E1*5B 1293G>C |
rs2031920 | (G>C) | FP: 5’-ACC CCA ATG GGT GTC TGT C-3’ RP: 5’-TCA TTC TGT CTT CTA ACT GGC AAT-3’ |
95°C for 5 min , 30 cycles of 95°C- 30 sec, 54°C- 45 sec, 72°C- 30 sec , final extension 72°C for 5 min |
576 bp | 1 Unit of PstI Incubation at 37˚C for 1h |
294 bp, 284 bp | 576 bp, 294 bp 282 bp |
576 bp |
|
CYP2E1*6 7632T>A |
rs6413432 | (T>A) | FP: 5’-AGG CTC GTC AGT TCC TGA AA -3’ RP: 5’-AAG GCA GGA GGA TGA CTT GA -3’ |
95°C for 5 min, 30 cycles of 95°C- 20 sec, 62°C- 25 sec, 72°C- 20 sec, final extension 72°C for 5 min |
685 bp | 1 Unit of DraI Incubation at 37˚C for 1h |
376 bp, 309 bp | 685 bp, 376 bp 309 bp |
685 bp |
|
CYP2E1*7B 71G>T |
rs6413420 | (G>T) | FP: 5’-CTG GAG TTC CCC GTT GTC TA-3’ RP: 5’-GGG TGA AGG ACT TGG GAA TA -3’ |
95°C for 5 min, 30 cycles of 95°C- 30 sec, 54°C- 45 sec, 72°C- 30 sec, final extension 72°C for 5 min |
547 bp | 1 Unit of DdeI Incubation at 37˚C for 1h |
301 bp, 246 bp | 547 bp, 301 bp 246 bp |
547 bp |