Table 2.

Primers and probes used for Leishmania qPCR diagnosis and species identification^a

Type of DNA targeted, primer set designation	SYBR green primer sequence		TaqMan primer sequenceb	Sequence sourcec	Source or reference
	Forward	Reverse
Minicircle
kDNA 1 minicircle	GGGTAGGGGCGTTCTGC	TACACCAACCCCCAGTTTGC		M94088	J.L.W.c
kDNA 2	AACTTTTCTGGTCCTCCGGGTAG	ACCCCCAGTTTCCCGCC		EU437406	Leish1 and -2 (20)
kDNA 3 minicircle	GGGTAGGGGCGTTCTGC	CCCGGCCTATTTTACACCAACC	M94088	J.L.W.
kDNA 4 minicircle	GGGTGCAGAAATCCCGTTCA	CCCGGCCCTATTTTACACCA	ACCCCCAGTTTCCCGCCCCG	Conserved kDNAd	J.L.W.
kDNA 5 minicircle	CTTTTCTGGTCCTCCGGGTAGG	CCACCCGGCCCTATTTTACACCAA	TTTTCGCAGAACGCCCCTACCCGC	L. infantum contig 1335	TaqMan qPCR (40)e
kDNA 7 minicircle	AATGGGTGCAGAAATCCCGTTC	CCACCACCCGGCCCTATTTTAC	CCCCAGTTTCCCGCCCCGGA	Conserved kDNAd	J.L.W.
L. (L.) amazonensis kDNA 1	GGTCCCGGCCCAAACTTTTC	CCGGGGTTTCGCACTCATTT		U19810	J.L.W.
L. (L.) amazonensis kDNA 2	GGTAGGGGCGTTCTGCGAAT	CCCGGCCTATTTTACACCAACC		EU370875	J.L.W.
L. (L.) amazonensis kDNA 3	GGGTAGGGGCGTTCTGC	TACACCAACCCCCAGTTTGC		M94089	J.L.W.
L. (L.) amazonensis kDNA 4	TGAGTGCAGAAACCCCGTTCATA	ACACCAACCCCCAGTTGTGA		EU370875	J.L.W.
L (V.) braziliensis kDNA 1	AATTTCGCAGAACGCCCCTAC	GTACTCCCCGACATGCCTCTG		U19807	J.L.W.
L (V.) braziliensis kDNA 3	TGCTATAAAATCGTACCACCCGACA	GAACGGGGTTTCTGTATGCCATTT	TTGCAGAACGCCCCTACCCAGAGGC	AF231100	Adapted from PCR (56)f
L (L.) infantum minicircle 1	TCCGCAGGAGACTTCGTATG	CACGACTATCCACCCCATCC	CTGAGAGACCCGCCGGGGCG	AF032997	Adapted from nested PCR (51)
L. (L.) major minicircle 1	ACGGGGTTTCTGCACCCATT	GTAGGGGCGTTCTGCGAAAA		LM15_BIN_Contig406	J.L.W.
L. (L.) mexicana minicircle 1	AATGCGAGTGTTGCCCTTTTG	GCCGAACAACGCCATATTAACC		AY145437	J.L.W.
L. (L.) tropica minicircle 1	GGGGGTTGGTGTAAAATAGGG	ACCACCAGCAGAAGGTCAAAG	TCCTGGCGGGGGTTTTCGCT	AF308689	J.L.W.
L. (L.) donovani minicircle 1	GCGGTGGCTGGTTTTAGATG	TCCAATGAAGCCAAGCCAGT	CCCATACCACCAAACGCAGCCCA	FJ416603	J.L.W.
Maxicircle
Cytochrome B 1	ATTTTAGTATGAGTGGTAGGTTTTGTT	CAATAACTGGGACGGTTGCT	CCATGTACGATGATGTCGTATTGAGGTCTAACA	AB095960	J.L.W.
L. (L.) amazonensis cytochrome B 1	GCGGAGAGGAAAGAAAAGGCTTA	AAAAGTCATGCTAAACACACACCACA		AB095964	J.L.W.
L. (L.) tropica cytochrome B 1	CAGGTTGCTTACTACGTGTTTATGGTG	TCGTATTACAAACCCTAAATCAAAATCTCA		AB095960	J.L.W.
L. (L.) tropica cytochrome B 2	TCAGGTTGCTTACTACGTGTTTATGGTG	TGCTAAACAAACACCACATATGATCTGC		AB095960	J.L.W.
L. (L.) tropica cytochrome B 3	TGACACACATATTTTAGTGTGGGTGGTAGG	TCCCCAATAAGACATCATTGTACATGGTAA		EF579904	J.L.W.
L. (L.) tropica cytochrome B 4	CACATATTTTAGTGTGGGTGGTAGGTTTTG	TCCCCAATAAGACATCATTGTACATGGTAA		EF579904	J.L.W.
Maxicircle 1	GCTTGGTTGGATTATTTTTGCTG	AACAACATTTTAACTCTTGTAGGATTCG	CTTTAGGTAGGGAGTTGTACTACGTTTTTTGACCT	DQ492252	J.L.W.
Genomic
Alpha-tubulin 1	GAGGTGTTTGCCCGCATC	CTCGCCCATGTCGTCG	TGAGGGCATGGAGGAGGGCG	XM_001681731	J.L.W.
DNA polymerase 1	TGTCGCTTGCAGACCAGATG	GCATCGCAGGTGTGAGCA	CAGCAACAACTTCGAGCCTGGCACC	AF009147	TaqMan qPCR (8)e
DNA polymerase 2	AGGAGGATGGCAAGCGGAAG	GCGACGGGTACAGGGAGTTG	TGGGGTCGAGCACCATGCCGCC	AF009136	J.L.W.
Miniexon 1	CGAAACTTCCGGAACCTGTCTT	CACCACACGCACGCACAC	CGGCAAGATTTTGGAAGCGCGCA	AL389894	J.L.W.
Miniexon 2	GTGTGGTGGCGGGTGTATGT	GCCCAGGTCGCTGTGAGG		LbrM02_V2.0710	J.L.W.
MAG 1 (MSP-associated gene 1)	AGAGCGTGCCTTGGATTGTG	CGCTGCGTTGATTGCGTTG	TGCGCACTGCACTGTCGCCCCC	AF058760	J.L.W.
MAG 2 (MSP-associated gene 2)	AGTTTTGGTTGGCGCTCCTG	CCCACTCGCTTTCCTTGGTC	CGCTGAGAGCGAGGCAGGCACGC	AF058760	J.L.W.
SIDER repeat 1	CGACCCTGTCACCACCACAG	GAGGCCACCCTATCGCTGAC	AM937229	J.L.W.
L. (V.) braziliensis DNA polymerase 1	TCGTTGAGGGAGGAGGTGTTTC	TCGGCTTTGAGGTTGGCTTC		XM_001563712	J.L.W.
L. (V.) braziliensis DNA polymerase 2	ACGTCGCCAACTGCTTCACC	GTGTTCGCACCGCCTTGAC		XM_001563712	J.L.W.
L. major MAG 1	GTCGTTGTCCGTGTCGCTGT	CGCTGTGTGTGTCCGTGTGT		XM_001681328	J.L.W.
L (L.) amazonensis DNA polymerase 1	GACGACGACGAGGAGGATGG	GCGACGGGTACAGGGAGTTG		AF009136	J.L.W.
GPI	CCAGATGCCGACCAAAGC	CGCGCACGTGATGGATAAC		AM117195	(66)
HSP70-1	GAAGGTGCAGTCCCTCGTGT	CCTCCGTCTGCTTGCTCTTG		FN395029	J.L.W.
HSP70-4	TCGAGATCGACGCGTTGTT	CCGCACAGCTCCTCGAA		FN395037	J.L.W.
SLACS	GGAGAAACTCACGGCACAGG	GCGCCTCGTAGGTCACAGTT		XM_001562078	J.L.W.
Leptomonas DNA
Leptomonas miniexon 1	TGGAGCGGGTGCATTAACTC	GGTCTCGAGGTGCCCATGAC		S78663	J.L.W.
Leptomonas GAPDH 2	AGAAGCCGGATGTGCTTGTG	GCCCTCAGCCTTCACCTTGT		AF053738	J.L.W.
Human DNA
Human TNF-α 1	GCCCTGTGAGGAGGACGAAC	AAGAGGTTGAGGGTGTCTGAAGGA	CCTTCCCAAACGCCTCCCCTGCCCC	NM_000594	J.L.W.
Human TNF-α 2	GCGCTCCCCAAGAAGACAGG	TGCCACGATCAGGAAGGAGAAG	CACCGCCTGGAGCCCTGGGGC	NM_000594	J.L.W.
Human GAPDH 1	GGGCTCTCCAGAACATCATCC	CCAGTGAGCTTCCCGTTCAG		NG_007073	J.L.W.
Human GAPDH 2	CATCAAGAAGGTGGTGAAGCAG	CGTCAAAGGTGGAGGAGTGG		NG_007073	J.L.W.
Human GAPDH 3	GCATGGCCTTCCGTGTCC	CGCCTGCTTCACCACCTTCT		NG_007073	J.L.W.

^aAll sequences were developed for use in SYBR green and/or TaqMan assays.

^bThe TaqMan probe used for human DNA was 5′VIC/3′TAMRA. The TaqMan probe used for all other DNA was 5′FAM/3′TAMRA.

^cPrimers and probes were designed by one of us (J. L. Weirather [J.L.W.]) from sequences available on the World Wide Web. GenBank accession numbers are listed.

^dThe “conserved kDNA” sequences used multiple sources aligned with CLUSTALW.

^eFor TaqMan qPCR, the primers and probes are the exact ones used for TaqMan but not SYBR green assay reported in the literature cited.

^f“Adapted from” indicates that the targeted gene in the literature cited was used by J. L. Weirather to redesign primers and probes.