TABLE 2.
List of primers and probes used in the ITD 5.0 rRT-PCR assays by assay, primer and probe sequences, and binding regions
| Assay type and target | Primer or probe (dye)a | Oligonucleotide sequence (5′→3′) | nt positionb |
|---|---|---|---|
| Quadruplex | |||
| Sabin 1 | Sabin 1 2S | AGG TCA GAT GCT TGA AAG C | 2505–2523 |
| Sabin 1 probe A4 (Cy5) | CGC CCC CAC CGT TTC ACG GA | 2540–2559 | |
| Sabin 1 1A | CCA CTG GCT TCA GTG TTT | 2600–2583 | |
| Sabin 2 | Sabin 2 2S | CCG TTG AAG GGA TTA CTA AA | 2525–2544 |
| Sabin 2 probe (FAM) | ATT GGT TCC CCC GAC TTC CAC CAA T | 2550–2572 | |
| Sabin 2 1A | CGG CTT TGT GTC AGG CA | 2595–2579 | |
| Sabin 3 | Sabin 3 2S | AGG GCG CCC TAA CTT T | 2537–2552 |
| Sabin 3 probe (ROX) | TCA CTC CCG AAG CAA CAG | 2554–2571 | |
| Sabin 3 1A | TTA GTA TCA GGT AAG CTA TC | 2591–2572 | |
| PanEV (any EV) | PAN-EV S | GGC CCC TGA ATG CGG CTA ATC C | 458–480 |
| PAN-EV probe (VIC) | CCG ACT ACT TTG GGW GTC CGT GT | 546–568 | |
| PAN-EV A | GCG ATT GTC ACC ATW AGC AGY CA | 603–581 | |
| PanPV (any poliovirus) | PanPV/PCR-S1 | TTG GAG TTC TTC ACI TAI TCI MGI TTY GAY ATG | 2832–2864 |
| PanPV PCR-probe 1A (FAM) | TGR TTN ARI GCR TGI CCR TTR TT | 2926–2904 | |
| PanPV PCR-1A | GGA GCT CCG GGT GGG AYR TAC ATI ATY TGR TAI AC | 2962–2928 | |
| WPV1 (wild PV1) | WEAF WPV1 S | GTA CAA ACC AGT CAY GTN AT | 2661–2680 |
| SOAS WPV1 S | CGT ACA GAC TAG RCA YGT NAT | 2660–2680 | |
| WPV1 probe S (FAM) | CAT WAT GGT TAC RCA MGC ACC T | 2729–2750 | |
| WPV1 A | GAG AAT AAY TTG TCY TTK GAY GT | 2800–2778 | |
| PV2 (any serotype 2) | PV Type 2 S | GAT GCA AAY AAC GGI CAT GC | 2911–2930 |
| PV Type 2 probe S (FAM) | ATG ACT ATA CGT GGC AGA C | 2993–3011 | |
| PV Type 2 probe 1D S (FAM) | CRC CKA TIC CTG GYA | 2972–2986 | |
| PV Type 2 A | TCA TAA AAG TGG GAR TAC GCR TT | 3110–3088 | |
| PV Type 2 A 1C | TCG TAA AAA TGA GAA TAT GCA TT | 3110–3088 | |
| AFR WPV3 | |||
| WPV3-I | SOAS WPV3 S | CAG GGA GTA GAT GAY CTN AT | 2443–2462 |
| WEAF genotype | WEAF WPV3 S | CAG GGG GTT GAT GAY TTR AT | 2443–2462 |
| WPV3 probe S (Cy5) | CNC ARA ACA GYC TTC CGG ATA CC | 2504–2526 | |
| WPV3 A | ACK GTG TCT GAY GGN AC | 2623–2607 | |
| SOAS WPV3 | |||
| WPV3-II | SOAS 6S | GTY RTA CAR CGR CGY AGY AGR A | 2671–2692 |
| SOAS genotype | SOAS WPV3 probe S (FAM) | TTC TTY GCA AGI GGR GCR TGY GT | 2713–2735 |
| SOAS 5A | TCY TTR TAI GTR ATG CGC CAA G | 2816–2795 |
a
Probes labeled with 6-carboxyfluorescein (FAM) were quenched with black hole quencher 1 (BHQ1), those labeled with ROX were quenched with BHQ2, those labeled with Cy5 were quenched with BHQ3, and those labeled with VIC were quenched with 6-carboxytetramethylrhodamine (TAMRA).
b
All nucleotide positions are relative to those described by Toyoda et al. (34), except PanPV GenBank accession no. AY184219.1 and SOAS WPV3 GenBank accession no. AJ293918.1.