16S Methods Comparison, raw sequence reads

A prominent approach to studying the microbiome is the amplicon-based marker gene survey. Such PCR-based methods are susceptible to error and bias at many steps. Moreover, methods for preparing NGS amplicon libraries have proliferated and currently vary in many parameters. It stands to reason that biases introduced during PCR amplification will vary as a function of these parameters. In order to characterize these biases formally, we have compared several major categories of methods for amplicon-based NGS library preparation. Using mock community standards, we have systematically dissected the amplification process and provide mechanistic insight into the sources of experimental error and bias in amplicon-based microbiome experiments.