TABLE 1.
PCR primers and probes used in the real-time PCR assay
| Primer or probea | Nucleotide sequence (5′ → 3′) | Tm (°C) | Location within targetb | Gene detectedc |
|---|---|---|---|---|
| Cj-FI (forward) | TGCTAGTGAGGTTGCAAAAGAATT | 58.2 | 918-941 | hipO (100 bp) |
| Cj-RI (reverse) | TCATTTCGCAAAAAAATCCAAA | 60.9 | 1018-997 | |
| Cj-FAM probe | ACGATGATTAAATTCACAATTTTTTTCGCCAAA | 68.1 | 975-943 | |
| Cc-FI (forward) | CATATTGTAAAACCAAAGCTTATCGG | 58.3 | 331-357 | glyA (133 bp) |
| Cc-RI (reverse) | AGTCCAGCAATGTGTGCAATG | 58.2 | 464-444 | |
| Cc-VIC probe | TAAGCTCCAACTTCATCCGCAATCTCTCTCTAAATTT | 68.8 | 431-397 |
Primers and probes were designed by using the Primer Express program, version 2.0 (Applied Biosystems, Foster City, CA). The melting temperatures (Tm) of the primers ranged from 58 to 60°C, and those of the probes ranged from 68 to 70°C. The TaqMan probes were conjugated with fluorescent reporter dyes FAM 495 (C. jejuni-specific probe; Cj-FAM) or VIC 538 (C. coli-specific probe; Cc-VIC) at the 5′ ends and with the quencher dye TAMRA 555 at the 3′ ends (Applied Biosystems).
The positions of the oligonucleotides are listed relative to the initiation codons (+1 methionine) of the respective genes.
The nucleotide sequences were retrieved from the GenBank sequence database (http://www.ncbi.nlm.nih.gov) under accession numbers Z36940 (hipO) and AFI 36494 (glyA).