Table 2.
22 pairs of specific primers for SYBR Green I PCR.
| Primer set for duplex PCR | Species and subgroups | Target gene | PCR primers | GenBank accession no. | location | Product size (bp) | T m valuesa | References | |||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Name | Forward or revers | primers' sequences (5′ - 3′) | |||||||||
| First run | 1 Escherichia coli | eaeA | ∗b | eae | F2 | CATTGATCAGGATTTTTCTGGTGATA | Z11541 | 899-924 | 106 | 83.2 ± 0.2 | [14] |
| EPEC and EHEC | R | CTCATGCGGAAATAGCCGTTA | 1000-979 | ||||||||
| Staphylococcus aureus | femB | ∗ | FemB | fw | AATTAACGAAATGGGCAGAAACA | AF106850 | 277-299 | 93 | 80.8 ± 0.3 | [15] | |
| rv | TGCGCAACACCCTGAACTT | 370-351 | |||||||||
| 2 Campylobacter jejuni | C. jejuni- specific DNA | ∗ | AB | F | CTGAATTTGATACCTTAAGTGCAGC | AL111168 | 381135 | 86 | 79.1 ± 0.4 | [8] | |
| DNA | R | AGGCACGCCTAAACCTATAGCT | 381185 | ||||||||
| EAEC | astA | EAST-1 | S | GCCATCAACACAGTATATCC | L11241 | 63-82 | 106 | 84.9 ± 0.6 | [16] | ||
| AS | GAGTGACGGCTTTGTAGTCC | 168-148 | |||||||||
| 3 Vibrio parahaemoliticus | tdh | Tdh199 | F | GGTACTAAATGGCTGACATC | X54341 | 601-582 | 251 | 81.6 ± 0.3 | [17] | ||
| R | CCACTACCACTCTCATATGC | 351-370 | |||||||||
| Emetic Bacillus cereus | ces | ∗ | ces-TM | F | GATGTTTGCGACGATGCAA | DQ360825 | 8689-8707 | 65 | 80.4 ± 0.1 | This study | |
| R | CTTTCGGCGTGATACCCATT | 8793-8734 | |||||||||
| 4 Salmonella spp. | invA | ∗ | Styinva | JHO-2-right | TCGTCATTCCATTACCTACC | M90846 | 167-186 | 119 | 81.3 ± 0.4 | [5] | |
| JHO-3-left | AAACGTTGAAAAACTGAGGA | 285-234 | |||||||||
| Clostridium perfringens | cpe | GAP | 11 | GGTTCATTAATTGAAACTGGTG | X81849 | 583-604 | 154 | 78.3 ± 0.4 | [18] | ||
| 12 | AACGCCAATCATATAAATTACAGC | 712-736 | |||||||||
|
| |||||||||||
| Second and third runs | 5 ETEC (ST) | ST | STa | F | GCTAATGTTGGCAATTTTTATTTCTGTA | M25607 | 294-321 | 190 | 78.5 ± 0.2 | [19] | |
| R | AGGATTACAACAAAGTTCACAGCAGTAA | 483-456 | |||||||||
| Plesiomonas shigelloides | gyrB | PSG | 237-F | TTCCAGTACGAGATCCTGGCTAA | AJ300545 | 237-259 | 68 | 83.1 ± 0.2 | [13] | ||
| 304-R | TGAATCGACACGCCAGAGTTC | 304-284 | |||||||||
| 6 EAggEC | aggR | ∗ | aggR-Z | F | CAGAATCGTCAGCATCAGCTACA | Z18751 | 432-454 | 97 | 79.5 ± 0.3 | This study | |
| R | GATGCCCTGATGATAATATACGGAA | 358-382 | |||||||||
| EIEC & Shigella spp. | virA | virA | F | CTGCATTCTGGCAATCTCTTCACA | D26468 | 1589-1622 | 215 | 82.4 ± 0.3 | [20] | ||
| R | TGATGAGCTAACTTCGTAAGCCCTCC | 1813-1788 | |||||||||
| 7 Aeromonas hydrophila | ahh1 | ∗ | AHH1 | F | GCCGAGCGCCCAGAAGGTGAGTT | CP000462 | 1653360-82 | 133 | 89.8 ± 0.4 | [21] | |
| R | GAGCGGCTGGATGCGGTTGT | 1653492-73 | |||||||||
| ETEC (LT) | LT | LT | 1 | AGCAGGTTTCCCACCGGATCACCA | S60731 | 613-636 | 132 | 82.0 ± 0.3 | [22] | ||
| 2 | GTGCTCAGATTCTGGGTCTC | 744-725 | |||||||||
| 8 Providencia alcalifaciens | gyrB | PAG | 38F | TCTGCACGGTGTGGGTGTT | AJ300547 | 38-56 | 73 | 81.0 ± 0.2 | [13] | ||
| 110R | ACCGTCACGGCGGATTACT | 110-92 | |||||||||
| Enterotoxigenic B. cereus | nheB | ∗ | SG | F3 | GCACTTATGGCAGTATTTGCAGC | DQ153257 | 2101-2123 | 152 | 82.7 ± 0.4 | [23] | |
| R3 | GCATCTTTTAAGCCTTCTGGTC | 2252-2231 | |||||||||
| 9 Yersinia enterocolitica | yadA | ∗ | yadA-X | F | CCAGAACCAATTGCAATGCCT | X13882 | 1564-1543 | 100 | 81.6 ± 0.2 | This study | |
| Y. pseudotuberculosis | R | CTTTAAACAGCTTGTTCCAGCCA | 1465-1487 | 81.1 ± 0.3 | |||||||
| Campylobacter coli | ceuE | ∗ | CCceuE | 825F | ACGCGCACAAGGCATACTT | X88849 | 3513-3531 | 91 | 77.6 ± 0.3 | This study | |
| 915R | CCAGTATTCAGGATCAAGATAAATGATTT | 3603-3575 | |||||||||
| 10 Vibrio cholerae | hlyA | hlyA | 2272-F | AGCAGCGTGTGGGACAAGA | X51746 | 2272-2291 | 71 | 82.4 ± 0.1 | [13] | ||
| 2272-F | GCGGACCCTAATGCATCAAT | 2342-2323 | |||||||||
| Vibrio parahaemoliticus | trh | Trh | 250-F | GGCTCAAAATGGTTAAGCG | DQ359748 | 256-274 | 250 | 81.1 ± 0.1 | [17] | ||
| 251-R | CATTTCCGCTCTCATATGC | 505-487 | |||||||||
| Singl PCR | EHEC (Stx 1) | Stx1 | JMS1 | F | GTCACAGTAACAAACCGTAACA | EF441598 | 509-488 | 95 | 81.1 ± 1.0 | [12] | |
| R | TCGTTGACTACTTCTTATCTGGA | 415-437 | |||||||||
| Singl PCR | EHEC (Stx 2) | Stx2 | JMS2 | F | CGACCCCTCTTGAACATA | EF441616 | 140-157 | 108 | 81.7 ± 0.3 | [12] | |
| R | GATAGACATCAAGCCCTCGT | 247-228 | |||||||||
a: Average ± standard deviation of Tm values of 10 tests: b: ∗:new selected or designed primer