Brian J. Mailloux Barnard College For the Sloan Deep Carbon Workshop May 16, 2008
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Transcript of Brian J. Mailloux Barnard College For the Sloan Deep Carbon Workshop May 16, 2008
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Subsurface Microbial Carbon Cycling: Rates and Processes
orRecovery and Characterization
of a Deep Microbial Ecosystem
Brian J. Mailloux
Barnard College
For the
Sloan Deep Carbon Workshop
May 16, 2008
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Talk overview
• Background
• Sampling Requirements
• Use of Carbon isotopes
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State of Knowledge
• Examining depths to 120°C
• Lower cell numbers at greater depth
• Lower diversity at greater depths
• Slow
• Hard to sample
Can we use carbon isotopes to understand rates and turnover times and in the future link to diversity?
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State of Knowledge
Low Diversity from a 2.825 km deep fault (Lin et al.,)
0.0
1.0
2.0
3.0
4.0
102 103 104 105 106 107 108Cells/ml or Cells/g
De
pth
(k
m)
10
Pfiffner et al. 2006
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Requirements of Subsurface SamplingConstraints
• CLEAN
• Molecular sample constraints?
• Sample Size-How large a sample do we need?
• Location-Where and how can we sample?
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Requirements of Subsurface SamplingMolecular Constraints
• PCR– Nanograms of DNA
• Metagenomes– 10’s to 100’s of micrograms of DNA– Amounts can be lower with whole
genome amplification
• Isotopes– 100’s of micrograms of DNA
• PLFA’s generally have smaller sample sizes than DNA
Kno
wle
dge
DNA
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Requirements of Subsurface SamplingSample Size
• 1011 cells. (0.25 mg of DNA)
• ROCK– 103 cells/g therefore need 108 grams!!
• WATER– 103 cells/ml therefore need 105 liters (10,000L)
• At 1 gpm≈2 days
• If you have flowing water you can get good samples!
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Requirements of Subsurface SamplingLocation
• Cores– Access to novel locations– Expensive and size limited
• Wells– Access to novel locations– Deep wells can be hard to sample
• Mines– Access to the subsurface– Locations limited– Can get clean samples– Can go back repeatedly and run experiments
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Carbon Isotopes of DNA
• Bangladesh Example
• How it could be used in the deep subsurface
• 12C=99%, 13C=1%, 14C=1ppt but t1/2=5730 yr
• Microarrays
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Analyzing 14C of DNA Bangladesh ExampleAtmospheric derived 14C
• Sampled ~2000 liters from a 180’ deep well.
• Extracted DNA ~150μg (Not trivial!)
• 14C DOC ~5700 yr bp
• 14C DIC ~6240 yr bp
• 14C DNA ~300 yr bp
Small, Young, Labile Pool of Organic Carbon!
E. Reichert, Senior Thesis
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How can we use Carbon Isotopes to Understand Subsurface Growth
Rates?
14C is generated in situthrough decay of U and Th.
14C in DIC, Hydrocarbons, CH4…..
14C in Microbes (DNA)
Steady-stateProduction=Decay.
Steady-stateProduction=Decay.
No ProductionOnly Decay after incorporation!
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Imagining an Experiment
• Collect 14C and 13C of DNA, DIC, DOC and compound specific electron donors
• 14C of DNA should be “older” with a more negative Δ14C
• The Δ14C offset should be directly related to the turnover rate (“age”) of the microbes.
• Can then directly get to turnover times in the deep subsurface.
• Can then use a 14C microarray in a subsurface Beta Cage to relate specific genes to Δ14C
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Goals-Need to Link
• Isotopes
• Geochemistry
• Genomics/Proteomics
• With good subsurface access
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ConclusionsACKNOWLEDGEMENTS
T.C. Onstott and collaborators within his lab including: Dylan Chivian, Eric J. Alm, Eoin L. Brodie, David E. Culley, Thomas Gihring, Alla Lapidus, Li-Hung Lin, Steve Lowry, Duane P. Moser, Paul Richardson, Gordon Southam, Greg Wanger, Lisa M. Pratt, Adam P. Arkin, Terry C. Hazen, Fred J. Brockman, Duane Moser
Columbia University- Greg Freyer, Martin Stute, Lex van Geen, Elizabeth Reichert
LLNL-Bruce Buccholz