Samples were assayed by TaqMan PCR using three triplex sets (Aeromonas, Vibrio,
Photobacterium) and one positive control (TaqMan Exogenous Internal Positive
Control reagent, IPC; Applied Biosystems) per sample. The 25 mL PCR mixtures also
contained 1 TaqMan Gene Expression Master Mix (Applied Biosystems), 10 pM
primers, 5 pM probes and 1 mL (100 ng) of bacterial genomic DNA template (or IPC).
Amplification proceeded using a 7500 Real Time PCR system (Applied Biosystems)
at 50 8C for 2 min and 95 8C for 10 min followed by 40 cycles of 95 8C for 15 s and
58 8C for 1 min.