Optimization of the 16S rRNA sequencing analysis pipeline for studying in vitro communities of gut commensals
Arianna I. Celis, Andrés Aranda-Díaz, Rebecca N. Culver, Katherine S. Xue, David A. Relman, Handuo Shi, Kerwyn Casey Huang
Abstract
samples that optimizes financial cost, time/effort, and accuracy/reproducibility has yet to be established. Here, we systematically identify protocol elements that have significant impact, introduce bias, and/or can be simplified. Our results indicate that community diversity and composition are generally unaffected by substantial protocol streamlining. Additionally, we demonstrate that a strictly aerobic halophile is an effective spike-in for estimating absolute abundances in communities of anaerobic gut commensals. This time- and money-saving protocol should accelerate discovery by increasing 16S rRNA data reliability and comparability and through the incorporation of absolute abundance estimates.