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Table 2 The oligonucleotide primers and the Multiplex PCR programs used for amplification of virulence genes of E. coli isolates

From: Biofilm formation, antimicrobial susceptibility, serogroups and virulence genes of uropathogenic E. coli isolated from clinical samples in Iran

Gene

Primer name

Primer sequence (5'-3)

Size of product (bp)

PCR program

M-PCR volume (50 μL)

pap

pap3

pap4

GCAACAGCAACGCTGGTTGCATCAT

AGAGAGAGCCACTCTTATACGGACA

336

1 cycle:

94 °C ------------ 1 min.

30 cycle:

94 °C ------------ 60 s

63 °C ------------ 30 s

72 °C ------------ 90 s

1 cycle:

72 °C ------------ 5 min

5 μL PCR buffer 10X

1.25 mM Mgcl2

150 μM dNTP (Fermentas)

1 μM of each primers F & R

1.2 U Taq DNA polymerase (Fermentas)

3 μL DNA template

Sfa

sfa1

sfa2

CTCCGGAGAACTGGGTGCATCTTAC

CGGAGGAGTAATTACAAACCTGGCA

410

Afa

afa1

afa2

GCTGGGCAGCAAACTGATAACTCTC

CATCAAGCTGTTTGTTCGTCCGCCG

750

fimH

FimH1

FimH2

GAGAAGAGGTTTGATTTAACTTATTG

AGAGCCGCTGTAGAACTGAGG

559

1 cycle:

94 °C ------------ 3 min.

40 cycle:

94 °C ------------ 60 s

58 °C ------------ 70 s

72 °C ------------ 70 s

1 cycle:

72 °C ------------ 6 min

5 μL PCR buffer 10X

2 mM Mgcl2

200 μM dNTP (Fermentas)

0.4 μM of each primers F & R

1 U Taq DNA polymerase (Fermentas)

3 μL DNA template