Metagenomics: DNA sequencing of environmental samples

Key Points

  • DNA sequencing can provide insights into organisms that are difficult to study because they are inaccessible by conventional methods such as laboratory culture.

  • Isolation and sequencing of DNA from mixed communities of organisms (metagenomics) has revealed surprising insights into diversity and evolution.

  • Genomic DNA sequence, and even complete genomes in some cases, has been generated from organisms that exist only in tight association with other organisms, including various obligate symbionts and pathogens, members of natural microbial consortia and an extinct cave bear.

  • Partially assembled or unassembled genomic sequence from complex microbial communities has revealed the existence of novel and environment-specific genes.

  • Advances in DNA sequencing technology and computational methods promise to accelerate progress in metagenomics.

Abstract

Although genomics has classically focused on pure, easy-to-obtain samples, such as microbes that grow readily in culture or large animals and plants, these organisms represent only a fraction of the living or once-living organisms of interest. Many species are difficult to study in isolation because they fail to grow in laboratory culture, depend on other organisms for critical processes, or have become extinct. Methods that are based on DNA sequencing circumvent these obstacles, as DNA can be isolated directly from living or dead cells in various contexts. Such methods have led to the emergence of a new field, which is referred to as metagenomics.

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Figure 1: Sequencing of ancient DNA.

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Acknowledgements

This work was carried out under the auspices of the US Department of Energy's Office of Science, Biological and Environmental Research Program and the by the University of California, Lawrence Livermore National Laboratory, Lawrence Berkeley National Laboratory and Los Alamos National Laboratory. S.G.T. was supported by a National Institutes of Health National Research Service Award Training and Fellowship grant. We would like to thank P. Hugenholtz and T. Woyke for helpful comments on the manuscript.

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Correspondence to Edward M. Rubin.

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FURTHER INFORMATION

ARB

Community Sequencing Program Sequencing Plans for 2005

Dog Genome Project

ERGO

EstimateS

GenDB

Joint Genome Institute Protocols web site

PRIAM

Ribosomal Database Project

Wolbachia Genome Project

Glossary

METHANOGENS

A group of hydrogen-consuming Archaea that generate methane by reduction of carbon dioxide.

BIOFILM

A layered aggregate of microorganisms.

NORMAL FILTRATION

A process in which particles that are above a certain size are removed from a fluid by forcing the solution through a membrane containing pores of a defined size.

TANGENTAL FLOW FILTRATION

A process in which a fluid is pumped tangentially along the surface of a porous membrane and an applied pressure forces some of the fluid, as well as dissolved particles of sufficiently small size, across the membrane.

GRAM-POSITIVE BACTERIA

Members of the phyla Actinobacteria and Firmicutes, which have a single membrane and a thick cell wall that is made of crosslinked peptidoglycan and therefore can be stained with the Gram staining procedure.

PHYLOGENETIC ANCHORING

A technique that involves screening large-insert libraries made from environmental DNA for clones that contain phylogenetic marker genes, and sequencing those clones in their entirety.

HORIZONTAL GENE TRANSFER

The transfer of genetic material between the genomes of two organisms that does not occur through parent–progeny routes.

AXENIC

A pure culture of a single species of microorganism.

DIFFERENTIAL LYSIS

A technique that uses conditions that will only lyse certain cells so that the DNA from those cells can be isolated from other cells in a community.

PULSED-FIELD GEL ELECTROPHORESIS

The use of pulsed electrical fields of alternating polarity to separate large fragments of DNA.

METAGENOMIC

A term used to describe techniques that characterize the genomes of whole communities of organisms rather than individual species.

WHOLE-GENOME SHOTGUN

An approach to genomic sequencing that involves breaking the DNA into small pieces and cloning them into vectors, followed by sequencing the clones at random.

DENSITY GRADIENT

This occurs in a solution in which the concentration of the solute is lowest at the top and gradually becomes more dense towards the bottom.

STABLE ISOTOPE PROBING

A technique that relies on the incorporation of a substrate that is enriched in a stable isotope, such as 13C, to identify microorganisms that can metabolize that substrate.

FLOW CYTOMETRY

A technique that measures the fluorescence of individual cells as they pass through a laser beam in an individual stream.

AFFINITY PURIFICATION

A technique for purifying cells or molecules that is based on specific binding to a protein or other molecule that has been immobilized on a solid substrate, such as beads or a column.

ISOTHERMAL STRAND DISPLACEMENT

A DNA amplification technique that uses rolling-circle amplification with φ29 DNA polymerase to generate large quantities of DNA without thermal cycling.

PYROSEQUENCING

A DNA sequencing technique that relies on detection of pyrophosphate release on nucleotide incorporation rather than chain termination with dideoxynucleotides.

CONTIG

A continuous stretch of DNA sequence that is assembled from multiple independent sequencing reads.

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Tringe, S., Rubin, E. Metagenomics: DNA sequencing of environmental samples. Nat Rev Genet 6, 805–814 (2005). https://doi.org/10.1038/nrg1709

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