Table 2.

Optimized multiplex PCR reactions targeting for 7 housekeeping gene alleles used in E. coli MLST analysis.

mpxPCR1 (3 genes)		mpxPCR1 (4 genes)
Component	Volume (µL)	Component	Volume (µL)
Phusion High-Fidelity PCR Master Mix (2×)	12.5	Phusion High-Fidelity PCR Master Mix (2×)	17.5
adk Forward Primer (10 µM)	0.25	icd Forward Primer (10 µM)	0.3
adk Reverse Primer (10 µM)	0.25	icd Reverse Primer (10 µM)	0.3
fumC Forward Primer (10 µM)	0.25	mdh Forward Primer (10 µM)	0.5
fumC Reverse Primer (10 µM)	0.25	mdh Reverse Primer (10 µM)	0.5
gyrB Forward Primer (10 µM)	0.25	purA Forward Primer (10 µM)	0.3
gyrB Reverse Primer (10 µM)	0.25	purA Reverse Primer (10 µM)	0.3
H2O	9.6	recA Forward Primer (10 µM)	0.5
gDNA template	1	recA Reverse Primer (10 µM)	0.5
Total	25	H2O	12.3
		gDNA template	2
		Total	35