Abstract
Amplification, sequencing and analysis of the 16S rRNA gene affords characterization of microbial community composition. As this tool has become more popular and projects have grown in size and scope, greater sample multiplexing is becoming necessary while maintaining high quality sequencing. Here, modifications to the Illumina HiSeq 2500 platform are described that afford greater multiplexing and 300 bp paired-end reads of higher quality than produced by the current Illumina MiSeq platform. To improve the feasibility and flexibility of this method, a 2-Step PCR amplification protocol is also described that allows for targeting of different amplicon regions, thus improving amplification success from low bacterial bioburden samples.
Copyright
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