Oligonucleotide primers
| . | Oligonucleotide sequence . | Nucleotide position* (exon number) . | Target size (bp) . | Competitor size (bp) . |
|---|---|---|---|---|
| 5′ GAPDH primer | 5′-AAGGCTGAGAACGGGAAGCTTGTCATCAAT-3′ | 180-210 (exon 4) | 500 | 710 |
| 3′ GAPDH primer | 5′-TTCCCGTTCAGCTCAGGGATGACCTTGCCC-3′ | 651-680 (exon 8) | ||
| 5′ PCNA primer | 5′-TCCATCCTCAAGAAGGTGTTGGAG-3′ | 27-50 (exon 1) | 664 | 782 |
| 3′ PCNA primer | 5′-CAGACATACTGAGTGTCACCGTTG-3′ | 668-691 (exon 5) | ||
| 5′ DNMT1 primer | 5′-ACCGCTTCTACTTCCTCGAGGCCTA-3′ | 3242-3266 (exon 28) | 335 | 460 |
| 3′ DNMT1 primer | 5′-GTTGCAGTCCTCTGTGAACACTGTGG-3′ | 3551-3576 (exon 30) | ||
| 5′ DNMT3A primer | 5′-CACACAGAAGCATATCCAGGAGTG-3′ | 2070-2093† | 551 | 668 |
| 3′ DNMT3A primer | 5′-AGTGGACTGGGAAACCAAATACCC-3′ | 2597-2620 | ||
| 5′ DNMT3B primer | 5′-AATGTGAATCCAGCCAGGAAAGGC-3′ | 1942-1965 (exon 17) | 190 | 331 |
| 3′ DNMT3B primer | 5′-ACTGGATTACACTCCAGGAACCGT-3′ | 2109-2132 (exons 18/19) | ||
| 5′ p15-M primer‡ | 5′-GCGTTCGTATTTTGCGGTT-3′ | 148 | ||
| 3′ p15-M primer | 5′-CGTACAATAACCGAACGACCGA-3′ | |||
| 5′ p15-U primer1-153 | 5′-TGTGATGTGTTTGTATTTTGTGGTT-3′ | 154 | ||
| 3′ p15-U primer | 5′-CCATACAATAACCAAACAACCAA-3′ |
| . | Oligonucleotide sequence . | Nucleotide position* (exon number) . | Target size (bp) . | Competitor size (bp) . |
|---|---|---|---|---|
| 5′ GAPDH primer | 5′-AAGGCTGAGAACGGGAAGCTTGTCATCAAT-3′ | 180-210 (exon 4) | 500 | 710 |
| 3′ GAPDH primer | 5′-TTCCCGTTCAGCTCAGGGATGACCTTGCCC-3′ | 651-680 (exon 8) | ||
| 5′ PCNA primer | 5′-TCCATCCTCAAGAAGGTGTTGGAG-3′ | 27-50 (exon 1) | 664 | 782 |
| 3′ PCNA primer | 5′-CAGACATACTGAGTGTCACCGTTG-3′ | 668-691 (exon 5) | ||
| 5′ DNMT1 primer | 5′-ACCGCTTCTACTTCCTCGAGGCCTA-3′ | 3242-3266 (exon 28) | 335 | 460 |
| 3′ DNMT1 primer | 5′-GTTGCAGTCCTCTGTGAACACTGTGG-3′ | 3551-3576 (exon 30) | ||
| 5′ DNMT3A primer | 5′-CACACAGAAGCATATCCAGGAGTG-3′ | 2070-2093† | 551 | 668 |
| 3′ DNMT3A primer | 5′-AGTGGACTGGGAAACCAAATACCC-3′ | 2597-2620 | ||
| 5′ DNMT3B primer | 5′-AATGTGAATCCAGCCAGGAAAGGC-3′ | 1942-1965 (exon 17) | 190 | 331 |
| 3′ DNMT3B primer | 5′-ACTGGATTACACTCCAGGAACCGT-3′ | 2109-2132 (exons 18/19) | ||
| 5′ p15-M primer‡ | 5′-GCGTTCGTATTTTGCGGTT-3′ | 148 | ||
| 3′ p15-M primer | 5′-CGTACAATAACCGAACGACCGA-3′ | |||
| 5′ p15-U primer1-153 | 5′-TGTGATGTGTTTGTATTTTGTGGTT-3′ | 154 | ||
| 3′ p15-U primer | 5′-CCATACAATAACCAAACAACCAA-3′ |
The 5′ and 3′ primers of a pair for standard and competitive RT-PCR were designed for sequences in different exons so that they amplify only cDNA and not genomic DNA. Target size refers to the size of the PCR products from the gene transcripts and competitor size refers to the size of the PCR products from the competitor DNA. For methylation-specific PCR, primers designed by Herman and coworkers39 were used.
The nucleotide sequence positions are relative to the ATG start.
The exon-intron organization has not been determined.
M primer, a primer specific for methylated DNA.
U primer, a primer specific for unmethylated DNA.