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Volume 10, Number 10—October 2004

Escherichia coli and Community-acquired Gastroenteritis, Melbourne, Australia

Roy M. Robins-Browne*†Comments to Author , Anne-Marie Bordun*†, Marija Tauschek*, Vicki R. Bennett-Wood*†, Jacinta Russell†, Frances Oppedisano†, Nicole A. Lister*, Karl A. Bettelheim*, Christopher K. Fairley*, Martha I. Sinclair‡, and Margaret E. Hellard‡1
Author affiliations: *University of Melbourne, Melbourne, Victoria, Australia; †Murdoch Childrens Research Institute, Melbourne, Victoria, Australia; ‡Monash University, Melbourne, Victoria, Australia

Main Article

Table 3

Characteristics of the polymerase chain reaction (PCR) primers used in this study for strain characterization

Primer Primer sequence (5′ to 3′) Target PCR program (30 cycles)a Product 
size (bp) Reference
P2 CCAGACGATACGATCCAG eaec 94°C, 30 s; 53°C, 30 s; 72°C, 60 s 917 (17)
P3 CTGGAGTTGTCGATGTT eae-α 94°C, 30 s; 53°C, 30 s; 72°C, 120 s 1,648 (17)
P4 GTAATTGTGGCACTCC eae-β 94°C, 30 s; 53°C, 30 s; 72°C, 120 s 1,926 (17)
P5 GCCTCTGACATTGTTAC eae-γ 94°C, 30 s; 53°C, 30 s; 72°C, 120 s 1,770 (17)
ecsD-lower TATTTTCAAAAAGAATGATGTC eae 94°C, 30 s; 56°C, 60 s; 72°C, 150 s ≈2,990d (18)
LP5 AGCTCACTCGTAGATGACGGCAAGCG eae-ε 94°C, 30 s; 55°C, 60 s; 72°C, 120 s 2,608 (18)
LP6B TAGTTGTACTCCCCTTATCC eae-ζ 94°C, 30 s; 53°C, 60 s; 72°C, 150 s 2,430 (18)
LP7 TTTATCCTGCTCCGTTTGCT eae-ι 94°C, 30 s; 52°C, 60 s; 72°C, 150 s 2,685 (18)
LP8 TAGATGACGGTAAGCGAC eae-η 94°C, 30 s; 52°C, 60 s; 72°C, 150 s 2,590 (18)
LP10 GGCATTGTTATCTGTTGTCT eae-κ 94°C, 30 s; 52°C, 60 s; 72°C, 150 s 2,769 (18)
LP11B GTTGATAACTCCTGATATTTTA eae-θ 94°C, 30 s; 50°C, 60 s; 72°C, 150 s 2,686 (18)
AGCAGGCATAACGCAAG lpfD 94°C, 60 s; 48°C, 50 s; 72°C, 60 s 798 (19)
AGTGCCCGTGTTCTTGAACTG efa1 94°C, 30 s; 50°C, 30 s; 72°C, 120 s 2,226 (20)
CGCGAGTGACGGCTTTGTAG astA 94°C, 30 s; 50°C, 60 s; 72°C, 90 s 109 (10)
ACAGAATCGTCAGCATCAGC aggR 94°C, 30 s; 50°C, 60 s; 72°C, 45 sf 254 (21)

aBefore the first cycle, the sample was denatured for 10 min at 95°C; after the last cycle, the sample was extended for 7 min at 72°C.
bPrimer P1 was used as forward primer in all PCR reactions in combination with P2, P3, P4, and escD-lower.
cConserved region of eae.
dAmplicon sequenced.
ePrimer SK1 was used as forward primer in all PCR reactions in combination with LP5, LP6B, LP7, LP8, LP10, and LP11B.
f35 cycles.

Main Article

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1Current affiliation: Burnet Institute, Melbourne, Victoria, Australia.

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