Table 1.
Primers for the strand-specific PCR and PCR conditions
| Aim | Primers | Primer sequence 1 | PCR condition |
|---|---|---|---|
| First strand synthesis of DHFR transcribed strand | DHFR_UP-TSP1 | 5'-GAGTCGAGTCGACATCGAGCCACTGTGCCCGGTCAGAC-3' | 94°C 15 s, 62°C 15 s, 72°C 60 s for 1 cycle |
| First strand synthesis of DHFR non-transcribed strand | DHFR_UP-NTSP1 | 5'-GAGTCGAGTCGACATCGGGATGTGCCCTGGTGACTGGA-3' | 94°C 15 s, 62°C 15 s, 72°C 60 s for 1 cycle |
| Amplification of DHFR transcribed strand | DHFR_TSP2 | 5'-GGATGTGCCCTGGTGACTGGA-3' | 94°C 15 s, 62°C 15 s, 72°C 40 s for 25 cycles |
| UP | 5'-GAGTCGAGTCGACATCG-3' | ||
| Amplification of DHFR no-transcribed strand | DHFR_NTSP2 | 5'-AGCCACTGTGCCCGGTCAGAC-3' | 94°C 15 s, 62°C 15 s, 72°C 40 s for 25 cycles. |
| UP | 5'-GAGTCGAGTCGACATCG-3' | ||
| First strand synthesis of c-myc transcribed strand | c-myc_UP-TSP1 | 5'-GAGTCGAGTCGACATCGAACTTTGTGCCTTGGATTT-3' | 94°C 15 s, 58°C 15 s, 72°C 60 s for 1 cycle |
| Amplification of c-myc transcribed strand | c-myc_TSP2 | 5'-TACTGGAGCTGATGCTGAGC-3' | 94°C 15 s, 58°C 15 s, 72°C 40 s for 25 cycles |
| UP | 5'-GAGTCGAGTCGACATCG-3' | ||
| Amplification of β-actin | β-actin P1 | 5'-GCCAGGTCCAGACGCAGGAT-3' | 94°C 15 s, 62°C 15 s, 72°C 10 s for 22 cycles |
| β-actin P2 | 5'-TGCTATCCCTGTACGCCTCTG-3' | ||
1 The underline sequence is an adaptor universal primer (UP) without homology to mammalian genomic DNA.