TABLE 2.
PCR primers and DNA amplification conditions for 16S, gyrB, secA1, hsp65, and rpoB
| Target | Region | Forward primer (5′-3′) |
Reverse primer (5′-3′) |
PCR cycling conditions | Reference or source | ||
|---|---|---|---|---|---|---|---|
| Primer name (position) | Sequence | Primer name (position) | Sequence | ||||
| 16S | bp 31-492 | E8F (bp 11-30) | AGAGTTTGATCCTGGCTCAG | 534r (bp 493-509) | ATTACCGCGGCTGCTGG | 30 s 98°C; 35× 5 s 98°C, 5 s 56°C, 20 s 72°C; 1 min 72°C | 3, this study |
| gyrB | bp 1031-1511 | Noc-gyrB-F (bp 972-992) | CTTCGCCAACACCATCAACAC | Noc-gyrB-R (bp 1563-1582) | TGATGATCGACTGGACCTCG | 30 s 98°C; 35× 5 s 98°C, 5 s 60°C, 20 s 72°C; 1 min 72°C | This study |
| Noc-gyrB-F3 (bp 1011-1030) | CGAGGAGGGCTTCCGCGCGG | Noc-gyrB-R3 (bp 1512-1532) | ATCGACTGGACCTCGTTGTTC | 30 s 98°C; 35× 5 s 98°C, 5 s 60°C, 20 s 72°C; 1 min 72°C | This study | ||
| secA1 | bp 431-875 | secA1-F47 (bp 413-430) | GCGACGCCGAGTGGATGG | secA1-ConR2 (bp 876-896) | TTGGCCTTGATGGCGTTGTTC | 30 s 98°C; 35× 5 s 98°C, 5 s 67°C, 20 s 72°C; 1 min 72°C | This study |
| secA1-F47 (bp 413-430) | GCGACGCCGAGTGGATGG | secA1-ConR (bp 913-933) | GCGGACGATGTAGTCCTTGTC | 30 s 98°C; 35× 5 s 98°C, 5 s 63°C, 20 s 72°C; 1 min 72°C | 10, this study | ||
| hsp65 | bp 165-565 | Noc-hsp65-F (bp 145-164) | ACCAACGATGGTGTGTCCAT | Noc-hsp65-R (bp 566-585) | CTTGTCGAACCGCATACCCT | 30 s 98°C; 35× 5 s 98°C, 5 s 54°C, 20 s 72°C; 1 min 72°C | 37 |
| Noc-hsp65-F2 (bp 109-129) | GTTGTCCTGGAGAAGAAGTGG | Noc-hsp65-R (bp 566-585) | CTTGTCGAACCGCATACCCT | 30 s 98°C; 35× 5 s 98°C, 5 s 54°C, 20 s 72°C; 1 min 72°C | 37, this study | ||
| rpoB | bp 1089-1488 | Noc-rpoB-F2 (bp 897-919) | CCGCTACAAGATCAACAAGAAGC | Noc-rpoB-R4 (bp 1612-1634) | CCCGCGAGGACATCGTCG | 30 s 98°C; 35× 5 s 98°C, 5 s 50°C, 20 s 72°C; 1 min 72°C | This study |
| Noc-rpoB-F2 (bp 897-919) | CCGCTACAAGATCAACAAGAAGC | Noc-rpoB-R2 (bp 1657-1677) | GGCGACGTACTCCATCTCCTC | 30 s 98°C; 35× 5 s 98°C, 5 s 50°C, 20 s 72°C; 1 min 72°C | This study | ||
| Noc-rpoB-F4 (bp 962-979) | CCCGCGAGGACATCGTCG | Noc-rpoB-R4 (bp 1612-1634) | CCCGCGAGGACATCGTCG | 30 s 98°C; 35× 5 s 98°C, 5 s 50°C, 20 s 72°C; 1 min 72°C | This study | ||
| Noc-rpoB-F5 (bp 987-1006) | CGAGTACCTGGTGCGYCTGC | Noc-rpoB-R5 (bp 1588-1607) | TCGACCGGCGAGTTGGCCTG | 30 s 98°C; 35× 5 s 98°C, 5 s 62°C, 20 s 72°C; 1 min 72°C | This study | ||