Protocol outlines steps for extracting ample DNA from uncultured microbes

May 06, 2014
UBC researchers probed the hidden microbial riches of British Columbia’s pristine Sakinaw Lake, situated on the Sunshine Coast. The collected samples were part of the microbial dark matter study led by the DOE JGI and described in a 2013 Nature paper. Credit: UBC Science, Flickr CC BY-NC-SA 2.0

Characterizing uncultured microbes starts with isolating and sequencing enough DNA from an environmental sample that may be as small as a single cell.

The protocol has been successfully employed to amplify the genomes of 201 single from uncultured microbes in a microbial dark matter study that was published in a July 2013 issue of Nature.

Though microbiologists say that there are more microbes in, on and around the Earth than there are stars in the galaxy, only a tiny fraction of these bacteria and archaea are known. One of the challenges scientists face in studying them is that most microbes don't thrive outside of their natural environments. Two methods used to learn more about such microbes are metagenomics and single-cell genomics. In July 2013, a Nature study led by U.S. Department of Energy Joint Genome Institute (DOE JGI) researchers employed single-cell genomics to provide information on 201 previously uncultured collected from nine sampling sites around the world. The information generated shed insight on 29 "mostly uncharted" branches on the tree of life.

The technique employed to extract and amplify the DNA from single microbial cells for the microbial dark matter study is now detailed in a Nature Protocols report published ahead online April 10, 2014. The DOE JGI team describes the steps they employed from sample preservation and preparation, to single cell separation through fluorescence-activated cell sorting (FACS), to cell lysis and ultimately, whole genome amplification.

The team reported that the procedure takes four days and yields up to a microgram of DNA per single , an amount considered "suitable" for PCR amplification and shotgun sequencing. They also noted that though the protocol was applied to microbial cells, it can also be adapted for microbial eukaryotes and mammalian cells.

Explore further: Sponges that sponge off bacteria

More information: Rinke C et al. "Obtaining genomes from uncultivated environmental microorganisms using FACS-based single-cell genomics." Nat Protoc. 2014 May;9(5):1038-48. DOI: 10.1038/nprot.2014.067. Epub 2014 Apr 10.

add to favorites email to friend print save as pdf

Related Stories

Microbial genomes help propose phylum name

Jun 05, 2013

At the phylum level, the number and diversity of unknown microbes still far outnumber those being studied. Metagenomics and single-cell genomics are tools helping researchers learn more about the "biological ...

Sponges that sponge off bacteria

Apr 08, 2014

Medical compounds harvested from a marine sponge are actually produced by symbiotic bacteria living in the sponges.

Bigelow laboratory scientists doach to study marine microbes

May 21, 2007

In a paper published this month in the Proceedings of the National Academy of Sciences, Dr. Ramunas Stepanauskas and Dr. Michael Sieracki have proven a new method of identifying genetic codes of ocean microbes from a sing ...

Recommended for you

Color-coading gene sequences in human cells

20 hours ago

(Phys.org)—Is there a way to peer inside the nucleus of a living cell and see how the genes interact? After the completion of the Human Genome Project in 2001, researchers have focused on epigenetic factors, ...

The origins of polarized nervous systems

Mar 03, 2015

(Phys.org)—There is no mistaking the first action potential you ever fired. It was the one that blocked all the other sperm from stealing your egg. After that, your spikes only got more interesting. Waves ...

New fat cells created quickly, but they don't disappear

Mar 03, 2015

Once fat cells form, they might shrink during weight loss, but they do not disappear, a fact that has derailed many a diet. Yale researchers in the March 2 issue of the journal Nature Cell Biology descri ...

User comments : 0

Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.