Title: Ultra-high throughput multiplexing and sequencing of >500 bp amplicon regions on the Illumina HiSeq 2500 platform

Johanna B. Holm, Michael S. Humphrys, Courtney K. Robinson, Matthew L. Settles, Sandra Ott, Li Fu, Hongqiu Yang, Pawel Gajer, Xin He, Elias McComb, Patti E. Gravitt, Khalil G. Ghanem, Rebecca M. Brotman, Jacques Ravel

Research output: Contribution to journalArticlepeer-review

Abstract

Amplification, sequencing and analysis of the 16S rRNA gene affords characterization of microbial community composition. As this tool has become more popular and amplicon-sequencing applications have grown in the total number of samples, growth in sample multiplexing is becoming necessary while maintaining high sequence quality and sequencing depth. Here, modifications to the Illumina HiSeq 2500 platform are described which produce greater multiplexing capabilities 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.

Original languageEnglish (US)
JournalUnknown Journal
DOIs
StatePublished - Sep 14 2018

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)

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