TABLE 2.

Multiplex PCR conditions and control strains used for detection of antimicrobial resistance genes in E. coli isolates

PCRa	Gene	Primer	Primer sequence	Final primer concn (μM)	Annealing temp (°C)	Product size (bp)	Control strain
1	sul1	sul1-Fb	CGGCGTGGGCTACCTGAACG	0.2	66	433	AMR 130g
		sul1-Bb	GCCGATCGCGTGAAGTTCCG	0.2
1	sul2	sulII-Lc	CGGCATCGTCAACATAACCT	0.3	66	721	AMR 130g
		sulII-Rc	TGTGCGGATGAAGTCAGCTC	0.3
1	sul3	sul3-GKa-Fd	CAACGGAAGTGGGCGTTGTGGA	0.2	66	244	RL0044k
		sul3-GKa-Rd	GCTGCACCAATTCGCTGAACG	0.2
2	tet(A)	TetA-Lc	GGCGGTCTTCTTCATCATGC	0.1	63	502	R08g
		TetA-Rc	CGGCAGGCAGAGCAAGTAGA	0.1
2	tet(B)	TetBGK-F2m	CGCCCAGTGCTGTTGTTGTC	0.2	63	173	PB#11g
		TetBGK-R2m	CGCGTTGAGAAGCTGAGGTG	0.2
2	tet(C)	TetC-Lc	GCTGTAGGCATAGGCTTGGT	0.5	63	888	PB#2g
		TetC-Rc	GCCGGAAGCGAGAAGAATCA	0.5
3	aadA	4Fe	GTGGATGGCGGCCTGAAGCC	0.1	63	525	AMR 075g
		4Re	AATGCCCAGTCGGCAGCG	0.1
3	strA/strB	strA-Ff	ATGGTGGACCCTAAAACTCT	0.4	63	893	AMR 075g
		strB-Rf	CGTCTAGGATCGAGACAAAG	0.4
3	aac(3)IV	aac4-Lg	TGCTGGTCCACAGCTCCTTC	0.2	63	653	AMR 075g
		aac4-Rg	CGGATGCAGGAAGATCAA	0.2
4	aadB	aadB-Li	GAGGAGTTGGACTATGGATT	0.2	55	208	TN1409h
		aadB-Ri	CTTCATCGGCATAGTAAAAG	0.2
4	aphA1	aph(3′)-Ia Fh	ATGGGCTCGCGATAATGTC	0.4	55	600	AMR61g
		aph(3′)-Ia Rh	CTCACCGAGGCAGTTCCAT	0.4
4	aphA2	aphA2-Li	GATTGAACAAGATGGATTGC	0.1	55	347	AMR 20g
		aphA2-Ri	CCATGATGGATACTTTCTCG	0.1
5	blaTEM	GKTEMFd	TTAACTGGCGAACTACTTAC	0.2	55	247	TEM4676l
		GKTEMRd	GTCTATTTCGTTCATCCATA	0.2
5	blaSHV	SHV-Fj	AGGATTGACTGCCTTTTTG	0.4	55	393	SHV4339l
		SHV-Rj	ATTTGCTGATTTCGCTCG	0.4
5	blaCMY-2	CMYFd	GACAGCCTCTTTCTCCACA	0.2	55	1,000	R1414d
		CMYRd	TGGACACGAAGGCTACGTA	0.2

^aMultiplex PCR 1 were done using the following thermal cycling conditions: one cycle consisting of 15 min at 95°C, 30 cycles consisting of 1 min at 95°C, 1 min at 66°C, and 1 min at 72°C, and one cycle consisting of 10 min at 72°C. Multiplex PCR 2 and 3 were done using the following thermal cycling conditions: one cycle consisting of 15 min at 94°C, 30 cycles consisting of 1 min at 94°C, 1 min at 63°C, and 1 min at 72°C, and one cycle consisting of 10 min at 72°C. Multiplex PCR 4 and 5 were done using the following thermal cycling conditions: one cycle consisting of 15 min at 94°C, 30 cycles consisting of 1 min at 94°C, 1 min at 55°C, and 1 min at 72°C, and one cycle consisting of 10 min at 72°C.

^bSee reference 17.

^cSee reference 18.

^dThis study.

^eSee reference 20.

^fSee reference 39.

^gSee reference 3.

^hSee reference 22.

ⁱSee reference 41.

^jSee reference 6.

^kSee reference 27.

^lObtained from Mike Mulvey (Winnipeg, Manitoba).

^mSee reference 11.