Table 2.
Primer/probe sets and final concentrations used in the four-plex RT-qPCR method, as well as reference amplicons for each method as used in the SCREENED analysis.
| Primer/probe name | Sequence (5′ → 3′) | Target sequence | Amplicon size (bp) | Location on reference genomea | Final concentration (nM) | Ref. |
|---|---|---|---|---|---|---|
| Target: CHIKV | ||||||
| CHIKV-a-F CHIKV-a-R CHIKV-a-P |
TGATCCCGACTCAACCATCCT GGCAAACGCAGTGGTACTTCCT FAM-TCCGACATCATCCTCCTTGCTGGC-BHQ-1 |
Nsp1 | 83 | 241–323 | 600 600 300 |
Panning et al. (2008) |
| Reference amplicon: TGATCCCGACTCAACCATCCTGGATATTGGTAGTGCGCCAGCAAGGAGGATGATGTCGGACAGGAAGTACCACTGCGTTTGCC | ||||||
| CHIKV-b-F CHIKV-b-R CHIKV-b-P |
TCACTCCCTGTTGGACTTGATAGA TTGACGAACAGAGTTAGGAACATACC HEX-AGGTACGCGCTTCAAGTTCGGCG-BHQ-1 |
E1 | 126 | 6856–6981 | 800 800 400 |
Lanciotti et al. (2007) |
| Reference amplicon: TCACTCCCTGTTGGACTTGATAGAGGCTGCTTTCGGAGAGATTTCCAGCTGTCATCTACCGACAGGTACGCGCTTCAAGTTCGGCGCCATGATGAAATCTGGTATGTTCCTAACTCTGTTCGTCAA | ||||||
| Target: ZIKV | ||||||
| ZIKV-a-F ZIKV-a-R ZIKV-a-P |
TTGGTCATGATACTGCTGATTGC CCYTCCACAAAGTCCCTATTGC TEX-CGGCATACAGYATCAGGTGCATWG GAG-BHQ-2 |
M/A | 77 | 941–1017 | 600 600 300 |
Adapted from Lanciotti et al. (2008) |
| Reference amplicon: TTGGTCATGATACTGCTGATTGCCCCGGCATACAGTATCAGGTGCATTGGAGTCAGCAATAGAGACTTCGTGGAGGG | ||||||
| ZIKV-b-F ZIKV-b-R ZIKV-b-P |
YCGYTGCCCAACACAAG CCACYAAYGTTCTTTTGCAGACAT Cy5-AGCCTACCTTGACAAGCARTCAGACA CTCAA-BHQ-2 |
E | 77 | 1192–1268 | 1000 1000 500 |
Adapted from Lanciotti et al. (2008) |
| Reference amplicon: TCGTTGCCCAACACAAGGTGAAGCCTACCTTGACAAGCAATCAGACACTCAATATGTCTGCAAAAGAACATTAGTGG | ||||||