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PeerJ. 2017 Sep 19;5:e3837. doi: 10.7717/peerj.3837. eCollection 2017.

Evaluation of ddRADseq for reduced representation metagenome sequencing.

Author information

1
The ithree institute, University of Technology Sydney, Sydney, New South Wales, Australia.

Abstract

BACKGROUND:

Profiling of microbial communities via metagenomic shotgun sequencing has enabled researches to gain unprecedented insight into microbial community structure and the functional roles of community members. This study describes a method and basic analysis for a metagenomic adaptation of the double digest restriction site associated DNA sequencing (ddRADseq) protocol for reduced representation metagenome profiling.

METHODS:

This technique takes advantage of the sequence specificity of restriction endonucleases to construct an Illumina-compatible sequencing library containing DNA fragments that are between a pair of restriction sites located within close proximity. This results in a reduced sequencing library with coverage breadth that can be tuned by size selection. We assessed the performance of the metagenomic ddRADseq approach by applying the full method to human stool samples and generating sequence data.

RESULTS:

The ddRADseq data yields a similar estimate of community taxonomic profile as obtained from shotgun metagenome sequencing of the same human stool samples. No obvious bias with respect to genomic G + C content and the estimated relative species abundance was detected.

DISCUSSION:

Although ddRADseq does introduce some bias in taxonomic representation, the bias is likely to be small relative to DNA extraction bias. ddRADseq appears feasible and could have value as a tool for metagenome-wide association studies.

KEYWORDS:

Double digest restriction site associated DNA sequencing; Human gut microbial communities; Metagenome-wide association studies; ddRADseq

Conflict of interest statement

The authors declare there are no competing interests.

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