Higher abundance of Actinomyces spp. associated with spontaneous preterm birth in overweight mothers

The V3–V4 region of the bacterial 16S rDNA gene was amplified by PCR (95°C for 3 min, followed by 30 cycles at 95°C for 30 s, 55°C for 30 s, 72°C for 45 s, and a final extension at 72°C for 10 min) using primers F 5’-TCG TCG GCA GCG TCA GAT GTG TAT AAG AGA CAG CCT ACG GGN GGC WGC AG-3’ and R 5’-GTC TCG TGG GCT CGG AGA TGT GTA TAA GAG ACA GGA CTA CHV GGG TAT CTA ATC C-3’ [29].

The amplicon DNA predicted size was about 550 bp and be verified by FragmentAnzlyzer (Agilent Technologies Inc.). Amplicon DNA, therefore, would be purified by KAPA Pure Beads (Roche) according to manufacturer’s protocol. Library index PCR would perform by Illumina Nextera XT DNA Library Preparation Kit Set A.Illumina MiSeq sequencingAmplicons were purified using the KAPA Pure Beads DNA (Roche) according to the manufacturer's instructions and quantified using Qubit 4 fluorometer (Thermo Fisher Scientific).