Single-Cell Genomics of Microbial Dark Matter

Gezielte Zellsortierung in der Einzelzellgenomik

BIOspektrum ◽

10.1007/s12268-021-1569-5 ◽

2021 ◽

Vol 27 (3) ◽

pp. 274-276

Author(s):

Morgan S. Sobol ◽

Anne-Kristin Kaster

Keyword(s):

Dark Matter ◽

Single Cell ◽

Cell Sorting ◽

Genetic Information ◽

Fluorescent Labeling ◽

Single Cell Genomics ◽

Rare Biosphere ◽

Prokaryotic Genomes ◽

Selection Of

AbstractSingle cell genomics (SCG) can provide reliable context for assembled genome fragments on the level of individual prokaryotic genomes and has rapidly emerged as an essential complement to cultivation-based and metagenomics research approaches. Targeted cell sorting approaches, which enable the selection of specific taxa by fluorescent labeling, compatible with subsequent single cell genomics offers an opportunity to access genetic information from rare biosphere members which would have otherwise stayed hidden as microbial dark matter.

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Single-cell Genomics: Defining Microbiology's Dark Matter

BioTechniques ◽

10.2144/000113848 ◽

2012 ◽

Vol 52 (5) ◽

pp. 301-303 ◽

Cited By ~ 1

Author(s):

Jeffrey M. Perkel

Keyword(s):

Dark Matter ◽

Single Cell ◽

Single Cell Genomics

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Printing Microbial Dark Matter: Using Single Cell Dispensing and Genomics to Investigate the Patescibacteria/Candidate Phyla Radiation

Frontiers in Microbiology ◽

10.3389/fmicb.2021.635506 ◽

2021 ◽

Vol 12 ◽

Author(s):

Sandra Wiegand ◽

Hang T. Dam ◽

Julian Riba ◽

John Vollmers ◽

Anne-Kristin Kaster

Keyword(s):

Dark Matter ◽

Single Cell ◽

Cell Sorting ◽

Treatment Plant ◽

Rrna Gene ◽

Label Free ◽

Single Cell Genomics ◽

Sorting Technique ◽

Unseen Species ◽

Candidate Phyla Radiation

As of today, the majority of environmental microorganisms remain uncultured. They are therefore referred to as “microbial dark matter.” In the recent past, cultivation-independent methods like single-cell genomics (SCG) enabled the discovery of many previously unknown microorganisms, among them the Patescibacteria/Candidate Phyla Radiation (CPR). This approach was shown to be complementary to metagenomics, however, the development of additional and refined sorting techniques beyond the most commonly used fluorescence-activated cell sorting (FACS) is still desirable to enable additional downstream applications. Adding image information on the number and morphology of sorted cells would be beneficial, as would be minimizing cell stress caused by sorting conditions such as staining or pressure. Recently, a novel cell sorting technique has been developed, a microfluidic single-cell dispenser, which assesses the number and morphology of the cell in each droplet by automated light microscopic processing. Here, we report for the first time the successful application of the newly developed single-cell dispensing system for label-free isolation of individual bacteria from a complex sample retrieved from a wastewater treatment plant, demonstrating the potential of this technique for single cell genomics and other alternative downstream applications. Genome recovery success rated above 80% with this technique—out of 880 sorted cells 717 were successfully amplified. For 50.1% of these, analysis of the 16S rRNA gene was feasible and led to the sequencing of 50 sorted cells identified as Patescibacteria/CPR members. Subsequentially, 27 single amplified genomes (SAGs) of 15 novel and distinct Patescibacteria/CPR members, representing yet unseen species, genera and families could be captured and reconstructed. This phylogenetic distinctness of the recovered SAGs from available metagenome-assembled genomes (MAGs) is accompanied by the finding that these lineages—in whole or in part—have not been accessed by genome-resolved metagenomics of the same sample, thereby emphasizing the importance and opportunities of SCGs.

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