Sample Preparation ● Sampling stool samples used in the majority of studies) mucosal biopsy samples(contain large amount of host dna ● nformation recording Where, when, and under which conditions the sampling is made Other factors(freshness, storage) o All sequencing based technologies have an inherent extraction method bias(some microbes are orders of magnitude more difficult to lyse than others
Sample Preparation Sampling stool samples (used in the majority of studies) mucosal biopsy samples (contain large amount of host DNA) Information recording Where, when, and under which conditions the sampling is made Other factors (freshness, storage) All sequencing based technologies have an inherent extractionmethod bias (some microbes are orders of magnitude more difficult to lyse than others)
Sample Selection Representative microbial Region Microbe density, cells/mL clades Sequencing implications Small intestine Duodenum 103 Streptococcus, Lactobacillus Low bacterial biomass decreases contents Jejunum 10-10 Escherichia, Coryne bacterium DNa yields and increases leum107-10 difficulty of non-16S sequencing approaches Large intestine 10-10 12 Bacteroidetes. Firmicutes DNA and RNA in stool samples contents and stool Actinobacteria are almost completely Proteobacteria microbial (>99%); shotgun Methanobrevibacter. Candida DNA and RNA approaches are Mucosa Small intestine: TM7 Shotgun sequencing will yield Enterobacteriaceae large numbers of host reads: Pasteurellaceae joint host and microbial data Lachnospiraceae readily obtained Eubacterium, Streptococcus Large intestine: Bacteroidetes Firmicutes, Actinobacteria Fusobacteria. Akkermansia
Sample Selection