Microbial 16S rRNA gene metabarcoding

Course Overview

The 16s rRNA gene has become the standard marker for prokaryote phylogenetic analysis, and combined with high-throughput sequencing technologies it is widely used to infer the structure and composition of microbial communities. Due to the continuous improvements in sequencing technologies and bioinformatics tools, there is a wide choice of methods for sequencing and analysing 16S rRNA gene assemblies. This workshop is designed to give students the necessary background and practical experience of the strategies for the analysis of the diversity and structure of prokaryote communities, covering i) experimental design and primer choices; ii) wet-lab and library preparation options; iii) sequence quality control and analysis and iv) statistical analysis of microbial community data. The many sequencing and analysis options will be discussed, whereas a more in-depth tutorial using real sequence data will provide an opportunity for the student to practice 16S rRNA sequence analysis from raw sequence files to ecological interpretation. Course material, such as presentation slides and necessary model data, will be provided to the students.

Targeted audience

This workshop is intended for students and researchers interested in microbial ecology but who are not yet very familiar with the techniques involved. Choosing the appropriate primers, library preparation kits, sequencing methodologies and bioinformatics pipelines can be quite daunting to the uninitiated. This workshop will allow researchers interested more confidence in their methodology and analyses choices. The target audience include students of animal or plant microbiomes as well as those studying environmental microbial communities. It is assumed that the workshop attendees are interested in performing 16S rRNA metabarcoding using the Illumina MiSeq platform, although other sequencing technologies will be discussed during the workshop.

Teaching format

The workshop will consist of both lectures and practical classes. Background information will be provided to help workshop attendees choose the appropriate experimental design, primers, sequencing library preparation kits and to contextualise the bioinformatics and statistical analysis methods. Practical tutorials will be conducted on a step-by-step basis to guide the student from when receiving data from a sequence provider to obtaining plots and tables describing microbial community diversity, structure and relationships to environmental variables or host data.