First molecular evidence of bovine hemoplasma species (Mycoplasma spp.) in water buffalo and dairy cattle herds in Cuba

Table 4 Primers and probes used in this study for the real-time TaqMan PCR assays, and sequence analysis of M. wenyonii and “Ca. M. haemobos” from dairy cattle and buffalo

Target	Primer/Probe	Sequence (5'-3')	Amplicon length (bp)	Reference
Bovine GAPDH	GAPDH.463f	GGCGTGAACCACGAGAAGTATAA	120	Modified from Leutenegger et al. [33]
	GAPDH.582r	CCCTCCACGATGCCAAAGT
	GAPDH.489p	ATTO550-AYACCCTCAAGATTGTCAGCAATGCCTCCT-BHQ-2
M. wenyonii	MwenyoniiF	CCACGTGAACGATGAAGGTCTT	119	Modified from Meli et al. [14]
	MwenyoniiR	GGCACATAGTTAGCTGTCACTTATTCAA
	Mwen_P	6-FAM-AGTACCATCAAGGCGYGCTCATTTCCTAG-BHQ-1
	MycWen15f^a	ACACATGCAAGTCGAACGAG	1360	This study
	MycWen1374r^a	ATTGAATGTGGTTTTGACTAGTACTTT	1360	This study
“Ca. M. haemobos”	Mwen_short.forw	CCATGTGAACGATGAAGGTCTTT	90	Modified from Meli et al. [14]
	Mwen_short.rev	AGTTTGCTGTCACTTATTCATGAGGTA
	Mwen_short.p	YY-CTA¹TCA¹GTTRTTA¹TCCCTCA¹TAA-BHQ-1
	MHBforw^a	GAATTAATGCTGATGGTATGCCTAA	1393	Meli et al. [14]
	MHBrev^a	CCAATCAGAATGTTCACTCTAGATGC	1393	Meli et al. [14]

^aPrimers used for the sequencing reactions
Abbreviations: BHQ, black hole quencher; 6-FAM, 6-carboxyfluorescein; YY, Yakima yellow; A¹, 2-aminopurin: internal modification to increase melting temperature in substitution of the minor groove binder (used in the original publication)

ISSN: 1756-3305