Table-2.
Primers list, program and protocol of each primer.
| Primer | Program | Protocol | |||||
|---|---|---|---|---|---|---|---|
| All primers were synthesized by Sigma-Aldrich, USA | |||||||
| Initial denaturation | No. of cycles | Denaturation | Annealing | Extension | Final extension | ||
| (1) 16 S rRNA | - | 30 | 95°C for 1 min | 70°C for 45 s | 72°C for 1 min | 72°C for 10 min | (1) |
| (2) PVL | - | 30 | 94°C for 30s | 55°C for 30s | 72°C 1 min | 72°C for 10 min. | (1) |
| (3) CLFA | - | 35 | 94°C for 60 sec | 57°C for 60 sec | 72°C for 1 min | 72°C for 10 min. | (1) |
| (4) Coa | 95°C for 2 min | 30 | 95°C 30s | 58°C for 2 min | 72°C for 4 min | 72°C for 7 min | (1) |
| (5) Hla and (6) Hlb | 95°C for 5 min | 30 | 94°C for 60 sec | 55°C for 30 s | 72°C for 1 min | 72°C for 10 min. | (1) |
| (7) Enterotoxins | - | 25 | 94°C for 30 s | 50°C for 30 s | 72°C for 30 s | 72°C for 2 min | (2) |
| Protocol (1) | PCR was performed in a 50μl reaction mixture containing 2 μl of template DNA (approximately 500 ng/μl), 5 μl of ×10 PCR buffer (750 mM Tris–HCl (pH 8.8), 200 mM (NH4)2SO4, and 0.1% Tween 20), 200 μM of each of the four deoxynucleotide triphosphates, 1 U of Taq DNA polymerase (Roch Applied Science), and 50 pmol of each primert | ||||||
| Protocol (2) | PCR was performed in a 50μl reaction mixture containing 2 μl of template DNA (approximately 500 ng/μl), 5 μl of ×10 PCR buffer (750 mM Tris–HCl (pH 8.8), 200 mM (NH4)2SO4, and 0.1% Tween 20), 200 μM of each of the four deoxynucleotide triphosphates, 1 U of Taq DNA polymerase (Roch Applied Science), and 50 pmol of each primer | ||||||
Hlb=Betahemolysin, Hla=Alphahemolysin, Coa=Coagulase, CLFA=Clumping factor A, PVL: Panton valentine leukocidin, PCR: Polymerase chain reaction