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Updated: Feb 10, 2026

Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
Published on: August 29, 2014
Making and Sequencing Heavily Multiplexed, High-Throughput 16S Ribosomal RNA Gene Amplicon Libraries Using a
Ankur Naqib1, Silvana Poggi1, Weihua Wang1
1DNA Services Facility, Research Resources Center, University of Illinois at Chicago, Chicago, IL, USA.
Abstract:
Deep sequencing of polymerase chain reaction (PCR)-amplified small subunit (16S or 18S) ribosomal RNA (rRNA) genes fragments is commonly employed to characterize the composition and structure of microbial communities. Preparing genomic DNA for sequencing of such gene fragments on Illumina sequencers can be performed in a straightforward, two-stage PCR method, described herein. The protocol described allows for up to 384 samples to be sequenced simultaneously, and provides great flexibility in choice of primers.
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