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Table 8 PCR programs (temperature and volume) for detection of STEC O-serogroups, virulence factors and antibiotic resistance genes

From: Characterization of Escherichia coli virulence genes, pathotypes and antibiotic resistance properties in diarrheic calves in Iran

Gene PCR program PCR volume (50 μL)
O157, O145, O103, O26, O111 1 cycle:  
95°C ------------ 3 min. 5 μL PCR buffer 10X
30 cycle: 1.5 mM Mgcl2
95°C ------------ 20 s 200 μM dNTPe
58°C ------------ 40 s 0.5 μM of each primers F & R
72°C ------------ 30 s 1.25 U Taq DNA polymerasee
1 cycle: 2.5 μL DNA template
72°C ------------ 8 min  
O91, O128, O121, O113, O45 1 cycle:  
94°C ------------ 6 min. 5 μL PCR buffer 10X
34 cycle: 2 mM Mgcl2
95°C ------------ 50 s 150 μM dNTP
58°C ------------ 70 s 0.75 μM of each primers F & R
72°C ------------ 55 s 1.5 U Taq DNA polymerase
1 cycle: 3 μL DNA template
72°C ------------ 10 min  
LT, ST 1 cycle:  
94°C ------------ 6 min. 5 μL PCR buffer 10X
34 cycle: 1.5 mM Mgcl2
95°C ------------ 50 s 200 μM dNTP
56°C ------------ 70 s 0.5 μM of each primers F & R
72°C ------------ 50 s 1 U Taq DNA polymerase
1 cycle: 4 μL DNA template
72°C ------------ 6 min  
stx1, stx2, eae, hly, cnf1, cnf2, cdtIII, f17c 1 cycle:  
95°C ------------ 3 min. 5 μL PCR buffer 10X
34 cycle: 2 mM Mgcl2
94°C ------------ 60 s 200 μM dNTP
56°C ------------ 45 s 0.5 μM of each primers F & R
72°C ------------ 60 s 1.5 U Taq DNA polymerase
1 cycle: 5 μL DNA template
72°C ------------ 10 min  
aadA1, tetA, tetB, dfrA1, qnr, Sul1, aac[3]-IV, blaSHV, CITM, cat1, cmlA 1 cycle:  
94°C ------------ 8 min. 5 μL PCR buffer 10X
32 cycle: 2.5 mM Mgcl2
95°C ------------ 60 s 200 μM dNTP
55°C ------------ 70 s 0.5 μM of each primers F & R
72°C ------------ 2 min 2 U Taq DNA polymerase
1 cycle: 3 μL DNA template
  72°C ------------ 8 min  
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