Protocol detail

Fecal Microbiome Analysis in Allo-HCT Patients

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Fecal samples from a patient undergoing allo-HCT at Memorial Sloan Kettering Cancer Center (MSKCC) were collected following an institutional fecal biospecimen collection protocol (12 (link)). DNA extraction and 16S rRNA sequencing of fecal samples were performed as described previously (11 (link), 42 (link)). Briefly, genomic DNAs were extracted using phenol-chloroform/isoamyl alcohol and 0.1-mm zirconium beads and further purified using QIAamp Mini Spin Columns (Qiagen). The V4-V5 region of the 16S rRNA gene was PCR amplified and sequenced on the Illumina Miseq platform (2x250). The paired-end reads were analyzed using the UPARSE and MOTHUR pipelines (43 (link), 44 (link)). Total bacterial loads in each sample were estimated by 16S qPCR (please see supplemental methods, Text S2, posted at doi.org/10.6084/m9.figshare.7063823 for details).

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Jung H.J., Littmann E.R., Seok R., Leiner I.M., Taur Y., Peled J., van den Brink M., Ling L., Chen L., Kreiswirth B.N., Goodman A.L, & Pamer E.G. (2019). Genome-Wide Screening for Enteric Colonization Factors in Carbapenem-Resistant ST258 Klebsiella pneumoniae. mBio, 10(2), e02663-18.

Publication 2019

QIAamp Mini Spin Columns Miseq platform

16s rrna Cancer Chloroform Dnas Fecal Genomic Isoamyl alcohol Patient Phenol Rrna gene Zirconium

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Variable analysis

independent variables

Fecal sample collection protocol
DNA extraction method
16S rRNA sequencing approach

dependent variables

Bacterial composition and diversity in fecal samples
Total bacterial loads in fecal samples

control variables

Patients undergoing allo-HCT at Memorial Sloan Kettering Cancer Center (MSKCC)

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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