The complex microbiome from native semen to embryo culture environment in human in vitro fertilization procedure

Table 1 Specific primers and probes used for 454 pyrosequencing and qPCR

Study methods	Target groups (amplicon length, T_m, assay)	Primers/Probes	Sequence (5′-3′)	References
454 pyrosequencing	16S rRNA gene	TIBACB-r^a	TTGGCAGTCTCAG (NNNNNNNN)AGAGTTTGATCCTGGCTCAG	Fabrice et al. 2009 [11]
		TIBACA/TI357RA-f^a	GTCTCCGACTCAG (NNNNNNNN)CTGCTGCCTYCCGTA
		A	CCATCTCATCCCTGCGTGTCTCCGACTCAG
		B	CCTATCCCCTGTGTGCCTTGGCAGTCTCAG
Real-time PCR	Total bacteria (466 bp, 60 °C, TaqMan)	Univ-f Univ-r Univ (probe)	TGGAGCATGTGGTTTAATTCGA TGCGGGACTTAACCCAACA CACGAGCTGACGACA (AG)CCATGCA	Ott et al. 2004 [12]
	Enterobacteriaceae (195 bp, 58 °C, SYBR)	Eco1457-f Eco1652-r	CATTGACGTTACCCGCAGAAGAAGC CTCTACGAGACTCAAGCTTGC	Bartosch et al. 2004 [13]
	Staphylococcus sp. (560 bp, 62 °C, SYBR)	TstaG422 TstaG765	GGCCGTGTTGAACGTGGTCAAATCA TIACCATTTCAGTACCTTCTGGTAA	Martineau et al. 2001 [14]
	Corynebacterium sp. (516 bp, 60 °C, SYRR)	CF-1 CF-2	CGTAGGGTGCGAGCGTTGTCCG CGTTGCGGGACTTAACCCAACACT	Adderson et al. 2008 [15]

^aIn TIBACB and in TIBACA/TI357RA, A and B adapter sequences are underlined, the barcode is indicated by N-s in parentheses and specific primers’ sequences 8F and 357R are shown in italics. In reverse primer, the same barcode was incorporated to all samples

ISSN: 1477-7827