Reagent and laboratory contamination can critically impact sequence-based microbiome analyses.
BACKGROUND
The study of microbial communities has been revolutionised in recent years by the widespread adoption of culture independent analytical techniques such as 16S rRNA gene sequencing and metagenomics. One potential confounder of these sequence-based approaches is the presence of contamination in DNA extraction kits and other laboratory reagents.


RESULTS
In this study we demonstrate that contaminating DNA is ubiquitous in commonly used DNA extraction kits and other laboratory reagents, varies greatly in composition between different kits and kit batches, and that this contamination critically impacts results obtained from samples containing a low microbial biomass. Contamination impacts both PCR-based 16S rRNA gene surveys and shotgun metagenomics. We provide an extensive list of potential contaminating genera, and guidelines on how to mitigate the effects of contamination.


CONCLUSIONS
These results suggest that caution should be advised when applying sequence-based techniques to the study of microbiota present in low biomass environments. Concurrent sequencing of negative control samples is strongly advised.
Pathogen Genomics Group, Wellcome Trust Sanger Institute, Hinxton, UK. sb18@sanger.ac.uk.
Susannah J
SJ
Salter
Michael J
MJ
Cox
Elena M
EM
Turek
Szymon T
ST
Calus
William O
WO
Cookson
Miriam F
MF
Moffatt
Paul
P
Turner
Julian
J
Parkhill
Nicholas J
NJ
Loman
Alan W
AW
Walker
D059018:DNA Contamination; D007202:Indicators and Reagents / Q000032:analysis*; D007753:Laboratories; D056186:Metagenomics; D064307:Microbiota; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000032:analysis; D012475:Salmonella / Q000235:genetics*; D017422:Sequence Analysis, DNA
D007202:Indicators and Reagents; D012336:RNA, Ribosomal, 16S