Gut metagenome sequencing of sample 5P34
This study used genomic DNA extracted from fecal samples of eight pediatric patients (mean age, 9.9 years), who underwent allo-HSCT for high risk acute leukemia. Fecal samples were collected before HSCT and at different time points after the transplant, up to about 85 days post-HSCT, for a total of 32 samples. DNA libraries were prepared using Illumina TruSeq DNA PCR-Free Low throughput Library Preparation Kit, following the manufacturers instructions with few modifications. 100 nanograms of each genomic DNA was fragmented using a Covaris S2 sonication device in order to obtain 350 bp inserts. After sonication, DNA fragments were end-repaired and the appropriate library size was selected using different ratios of the Sample Purification Beads, then the samples were A-tailed and ligated to adapters. Because of the low qualities of the library obtained, an Enrich DNA Fragments step was introduced according to the manufactures instructions (Illumina TruSeq DNA Sample Preparation Kit). Finally, size and concentration of DNA fragments were assessed using KAPA Library Quantification Kits (Kapa Biosystems) specific for the Illumina libraries. Approximately 8 pM libraries were paired-end sequenced (2 x 100bp) on Illumina GAIIx platform: three samples were simultaneously sequenced in each line.