Table 1. PCRs and primers used in CoV detection*.
| PCRs (reference) | Primers, 5′→3′ | Nested fragment size, region (primer locations on the reference) genome)† | Type of CoV (no.) |
|---|---|---|---|
| Nested pan-CoV-I (6) | PLQ-F1, CGTTGGIACWAAYBTVCCWYTICARBTRGG | ≈400 nt, RdRp (18310–187450) | α-CoV (8), β-CoV (1) |
| PLQ-R1, GGTCATKATAGCRTCAVMASWWGCNACATG | |||
| PLQ-F2, GGCWCCWCCHGGNGARCAATT | |||
| PLQ-R2, GGWAWCCCCAYTGYTGWAYRTC | |||
| Nested pan-CoV-II (7) | WT-COV-F1, GGTTGGGAYTAYCCHAARTGTGA | ≈430 nt, RdRp (15260–15700) | α-CoV (5), β-CoV (2) |
| WT-COV-R1, CCATCATCASWYRAATCATCATA | |||
| WT-COV-F2, GAYTAYCCHAARTGTGAYAGAGC | |||
| WT-COV-F3, GAYTAYCCHAARTGTGAUMGWGC | |||
| Hemi-nested RdRp-sequence assay (9) | EMC-SeqRdRP-Rev, GCATWGCNCWGTCACACTTAGG | ≈230 nt, RdRp (15048–15290) | α-CoV (2), β-CoV (1) |
| EMC-SeqRdRP-Fwd, TGCTATWAGTGCTAAGAATAGRGC | |||
| EMC-SeqRdRP-Rnest, CACTTAGGRTARTCCCAWCCCA | |||
| Hemi-nested N-sequence assay (9) | EMC-SeqN-Fwd, CCTTCGGTACAGTGGAGCCA | ≈280 nt,N seq (29,549–29,860) | – |
| EMC-SeqN-Rev, GATGGGGTTGCCAAACACAAAC | |||
| EMC-SeqN-Fnest, TGACCCAAAGAATCCCAACTAC | |||
| Nested CII-pan-CoV-III | NM-CoV-2F1, ACWGTTCARGGICCWCCIGG | ≈355 nt, helicase (17,060–17,410) | β-CoV (2) |
| NM-CoV-2F2, GTTCARGGGCCWCCGGGNAC | |||
| NM-CoV-2R1, GGCAGCTGWGCWGGRTCICCNACRTA | |||
| NM-CoV-2R2, AGCTGWGCWGGRTCGCCIACRTANAC | |||
| Nested CII-MERS-RdRp | NM-HCOV-F1, GTGCTAAGAATAGAGCTCGCACT NM-HCOV-F2, AGAGCTCGCACTGTTGCAGGC | ≈190 nt, RdRp (15068–15249) | β-CoV (1, MERS CoV) |
| NM-HCOV-F2, AGAGCTCGCACTGTTGCAGGC | |||
| NM-HCOV-R1, ACCCATAAGATGCGGATTATCAAC | |||
| NM-HCOV-R2, TGCGGATTATCAACATCTTTGTAC | |||
| Hemi-nested CII-MERS N sequence | NM-NSeq-F-1, ACTTCCTTCGGTACAGTGGAGC | ≈170 nt, N seq (29545–29713) | – |
| NM-NSeq-R-1, GGCACTGTTCACTTGCAATC | |||
| NM-NSeq-R-2, GGAGGTTCAGACATTTGGTCT | |||
| upE and ORF1b real-time assays (8) | upE-Fwd: GCAACGCGCGATTCAGTT | Upstream of E gene and ORF 1b | – |
| upE-Prb: FAM-CTCTTCACATAATCGCCCCGAGCTCG-TAMRA | |||
| upE-Rev: GCCTCTACACGGGACCCATA | |||
| ORF1b-Fwd: TTCGATGTTGAGGGTGCTCAT | |||
| ORF1b-Prb: FAM-CCCGTAATGCATGTGGCACCAATGT-TAMRA | |||
| ORF1b-Rev: TCACACCAGTTGAAAATCCTAATTG |
* CoV, coronavirus; MERS, Middle East respiratory syndrome; RdRp, RNA-dependent RNA polymerase; –, not applicable; ORF, open reading frame. †Primer locations are based on human β-CoV 2c EMC/2012, complete genome (GenBank accession no. JX869059).