Comparison of Boiling and Robotics Automation Method in DNA Extraction for Metagenomic Sequencing of Human Oral Microbes

PLoS One. 2016 Apr 22;11(4):e0154389. doi: 10.1371/journal.pone.0154389. eCollection 2016.


The rapid improvement of next-generation sequencing performance now enables us to analyze huge sample sets with more than ten thousand specimens. However, DNA extraction can still be a limiting step in such metagenomic approaches. In this study, we analyzed human oral microbes to compare the performance of three DNA extraction methods: PowerSoil (a method widely used in this field), QIAsymphony (a robotics method), and a simple boiling method. Dental plaque was initially collected from three volunteers in the pilot study and then expanded to 12 volunteers in the follow-up study. Bacterial flora was estimated by sequencing the V4 region of 16S rRNA following species-level profiling. Our results indicate that the efficiency of PowerSoil and QIAsymphony was comparable to the boiling method. Therefore, the boiling method may be a promising alternative because of its simplicity, cost effectiveness, and short handling time. Moreover, this method was reliable for estimating bacterial species and could be used in the future to examine the correlation between oral flora and health status. Despite this, differences in the efficiency of DNA extraction for various bacterial species were observed among the three methods. Based on these findings, there is no "gold standard" for DNA extraction. In future, we suggest that the DNA extraction method should be selected on a case-by-case basis considering the aims and specimens of the study.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adsorption
  • DNA, Bacterial / genetics
  • DNA, Bacterial / isolation & purification*
  • Dental Plaque / microbiology*
  • High-Throughput Nucleotide Sequencing
  • Hot Temperature
  • Humans
  • Liquid Phase Microextraction / instrumentation
  • Liquid Phase Microextraction / methods*
  • Male
  • Metagenomics / instrumentation
  • Metagenomics / methods*
  • Microbiota / genetics
  • Pilot Projects
  • Principal Component Analysis
  • RNA, Ribosomal, 16S / genetics*
  • Robotics / methods*
  • Sequence Analysis, DNA


  • DNA, Bacterial
  • RNA, Ribosomal, 16S

Grant support

This work was supported by MEXT Tohoku Medical Megabank Project and was partially supported by the Center of Innovation Program from the Japan Science and Technology Agency (JST) and Grants-in-Aid from the Japan Society for the Promotion of Science to RY. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.