Table 2.

Primers used for PCR amplification and detection.

Primers	Sequences (5′-3′)a	Restriction enzyme	Amplification target
Div-F1	GGATCCATGGCACTTACAGCATT	BamHI	divIVA gene
Div-R1	CTCGAGTTCTTCTATTGAAAGTACTAC	XhoI	divIVA gene
STK-F1	GGATCCATGATTCAAATCGGTAAG	NdeI	stk gene
STK-R1	CTCGAGTTGTCCGCTACCTGTTG	HindIII	stk gene
Div-T172A-F1	CAACAGCAAGTTACATTCAAGCAAGTGACGAAG		Mutant divIVA gene
Div-T172A-F1	CTTGAATGTAACTTGCTGTTGGGCGGAGGAT		Mutant divIVA gene
Div-T199A-F1	AAAGTTTGGATTATACGCATCAATTGACACCAGAAG		Mutant divIVA gene
Div-T199A-F1	TTTCAAACCTAATATGCGTAGTTAACTGTGGTCTTC		Mutant divIVA gene
L1	CGAATTCGCTTTGCTAAGT TGGTTT	EcoRI	Upstream border of divIVA
L2	CGGATCCCTTTCCTCCTAAGTTTTAAC	BamHI	Upstream border of divIVA
Spc1	GGATCCGTTCGTGAATACATGTTATA	BamHI	SpcR gene
Spc2	GTCGACGTTTTCTAAAATCTGAT	SalI	SpcR gene
R1	CGTCGACATTTTAAGCGAGTAGGAG	SalI	Downstream border of divIVA
R2	GCATGCAGACTTGCTCAATAGGA	Sph1	Downstream border of divIVA
Check In1	GACGCAGATGAAGTTGATGACTT		Internal region of divIVA
Check In2	TGAATGTAACTTGCTGTTGGGC		Internal region of divIVA
Check Out1	ATATGTCGGAGCAACAAGCAAGAC		For combined PCR detection
Check Out2	TTCGATTTCAGCTTCTGCAAGG		For combined PCR detection

^aThe underlined sequences are the restriction sites or nucleotides involved in site directed mutation.