Rapid genetic typing of diarrheagenic Escherichia coli using a two-tube modified molecular beacon based multiplex real-time PCR assay and its clinical application

Table 4 The primers and probes used for multiplex real-time PCR assay in this study ^a F/R/P: forward and reverse primer, ^b concentration

Tube	Group	Target genes	Primers and Probes^a	Sequence(5′-3′)	Conc^b(μM)
Tube 1	ETEC¹	stp	stp- F	tag-AAAAGCGAGTGTACCTCGACA	0.18
			stp- R	tag-CAGTTGACTGACTAAAAGAGGGG	0.3
			stp- P	HEX-CGCGTCTCAAATATCCGTGAAACAACATGACGCG-Dabcyl	0.2
		sth	sth- F	tag-GTGGTCCTGAAAGCATGAATAG	0.12
			sth- R	tag-CAACAAAGCAACAGGTACATACG	0.12
			sth- P	FAM-CGCGGTGAATTGTGTTGTAATCCTGCTTGTACCGCG-Dabcyl	0.2
		lt	lt- F	tag-ACAGGAGGTTTCTGCGTTAG	0.18
			lt- R	tag-GGTGGGAAACCTGCTAATCT	0.18
			lt- P	ROX-CGCCGGTATTACAGAAATCTGAATATAGCTCCGGCG-Dabcyl	0.2
	EAEC¹	aggR	aggR- F	tag-TGCAAAAGAAGAAATCAACAGT	0.18
			aggR- R	tag-CAGAATCGTCAGCATCAGCTAC	0.3
			aggR- P	CY5-CGGACAAAAGTAGATGCTTGCAGTTGTCCG-Dabcyl	0.2
Tube 2	EIEC²	ipaH	ipaH- F3	tag-GAAAACCCTCCTGGTCCATC	0.32
			ipaH- R3	tag-GTCTGGAAGGCCAGGTAGACTT	0.32
			ipaH- P	FAM-CCCGGCTGGAGGACATTGCCCGGG-Dabcyl	0.2
	EPEC&	escV	escV- F	tag-GGCTCTCTTCTTCTTTATGGCTG	0.4
	EHEC²		escV- R	tag-GGGAAAGAAGTTAGTTCAAGAGGAT	0.4
			escV- P	HEX-CCCGCGCAACAGTTGTGGTGGATATCATTATCGCGGG-Dabcyl	0.2
		eaeA	eaeA- F	tag-GTAACCAGGCTTCGTCACA	0.32
			eaeA- R	tag-AAGGAAAAAACGCTGACCCG	0.32
			eaeA- P	CY5-CCCAGTGGTAATAACTTTGACGGTAGTTCACTGGG-Dabcyl	0.2
	EHEC²	stx1	stx1- F	tag-ASAGCGGTTACATTGTCTGGT	0.24
			stx1- R	tag-CTGCGTCAGTGAGGTTCCA	0.24
			stx1- P	ROX-CCGCGTACGGGGATGCAGATAAATCGCGG-Dabcyl	0.2
		stx2	stx2- F	tag-CATGACAACGGACAGCAGTTA	0.24
			stx2- R	tag-TCTGGTCATTGTATTACCACTGAA	0.24
			stx2- P	ROX-CCGCCACTCACTGGTTTCATCATATCTGGCGG-Dabcyl	0.2
Tube 1&2	IAC^1,2		IAC-F	GGCGCGCCTAACACATCT	0.1
			IAC-R	TGGAAGCAATGCCAAATGTGTA	0.1
			ICA-P	CGGTGGTTACAACGGGAGAAGACAATGCCACCG	0.2
			tag	GCAAGCCCTCACGTAGCGAA	2.4

ISSN: 1476-0711