GSM8880931: ok122_sequences.fastq.gz; Mus musculus; OTHER
DNA was extracted from mouse fecal samples using the PureLink Microbiome DNA Purification Kit (Invitrogen) and Mobio PowerSoil DNA Extraction Kit (Mo Bio). The V4 region of the bacterial 16S rRNA gene was amplified by polymerase chain reaction (PCR) using the 515F (AATGATACGGCGACCACCGAGATCTACACGCT) barcoded and 806R (TATGGTAATTGTGTGYCAGCMGCCGCGGTAA) primers. A reaction containing a final concentration of 0.04% of each primer and 0.5% of PrimeSTAR Max DNA Polymerase (TaKaRa) 50μl total volume. PCR reactions were carried out by 30-35 cycles of denaturation (95°C), annealing (55°C) and extension (72°C), with final elongation at 72°C. PCR products were purified using AMPure magnetic beads (Beckman Coulter) and quantified using a Denovix spectrophotometer. Samples were then pooled at equal amounts, loaded on 2% agarose E-Gel (Thermo Fisher), purified and sent for sequencing using the Illumina MiSeq platform (Genomic center, Azrieli Faculty of Medicine, BIU, Israel).