Metagenomic sequencing of stool from pediatric Ulcerative Colitis patient receiving Open_Label-_FMT
DNA was extracted from thawed stool samples using the DNeasy PowerSoil HTP 96 kit (Qiagen product number 12955-4). 16S rRNA gene libraries targeting the V4 region of the 16S rRNA gene were prepared by first normalizing template concentrations and determining optimal cycle number by way of qPCR.  Using primers 515F and 806rcbc0. To ensure minimal over-amplification, each sample was diluted to the lowest concentration sample, amplifying with this sample optimal cycle number for the library construction PCR using the Golay primer set (Caporaso, 2011 and 2012).  Four replicates were pooled, cleaned and normalized prior to sequencing on an Illumina MiSeq in paired-end mode (2x175bp) using custom index 5-ATTAGAWACCCBDGTAGTCC GG CTGACTGACT-3 and custom Read 1 (5-TATGGTAATT GT GTGYCAGCMGCCGCGGTAA-3) and Read 2 (5-AGTCAGTCAG CC GGACTACNVGGGTWTCTAAT-3) primers. 515F: 5-AATGATACGGCGACCACCGAGATCTACACTATGGTAATTGTGTGCCAGCMGCCGCGGTAA-3 806rcbc0: 5-CAAGCAGAAGACGGCATACGAGATTCCCTTGTCTCCAGTCAGTCAGCCGGACTACHVGGGTWTCTAAT-3