|
Primers used in methylation specific PCR and bisulfite genomic sequnecing |
||
| Methylation Specific PCR (MSP) |
Annealing |
positiona |
|
|
||
| Promoter A (methylated) |
58°C |
1711–1842 |
| Forward: 5'-TTC GTC GTA GTT TGC GAA TG |
1711–1730 |
|
| Reverse: 5'-CAA CAC GTA CCT AAA CGC GA |
1842-1821 |
|
| Promoter A (unmethylated) |
54°C |
1710–1843 |
| Forward: 5'-TTT GTT GTA GTT TGT GAA TGG |
1710–1730 |
|
| Reverse: 5'-ACA ACA CAT ACC TAA ACA CAA |
1843-1823 |
|
| Promoter B |
Ref. [23] |
|
|
|
||
| Real-time Quantitative MSP |
||
|
|
||
| Promoter B (Methylated) |
60°C |
1171–1293 |
| Forward: 5'-TTG TCG TAG CGT TCG GTT AC |
1171–1190 |
|
| Reverse: 5'-AAA CTA CAA CGC CCA CAA AA |
1293 – 1274 |
|
| Probe: 5'-Fam-TCG CGG GAC GGA TTC GTT TA |
1218–1237 |
|
|
|
||
| Bisulfite Genomic Sequencing |
||
|
|
||
| Promoter B |
58°C |
1141–1454 |
| Forward: 5'-tgg gtg gga ggg tgt tt |
1141–1157 |
|
| Reverse: 5'-cct cac aaa acc cta caa acc t |
1454-1433 |
|
|
a: Numbering is based on NCBI accession number U92868. | ||
Yang et al. BMC Cancer 2008 8:124 doi:10.1186/1471-2407-8-124 |
||