Jason dives 243 and 246 (18.47 N, 155.18 W).
URL | https://www.bco-dmo.org/dataset/616326 |
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Download URL | https://www.bco-dmo.org/dataset/616326/data/download |
Media Type | text/tab-separated-values |
Created | October 23, 2015 |
Modified | August 19, 2016 |
State | Final no updates expected |
Brief Description | Assembled metagenome sequences > 500 bp from EPR, Lo'ihi, and control |
Acquisition Description
Methodology:
EPR: Rock material was collected from the EPR at 9.725 N, 104.16 W (2,674 m depth) aboard the R/V Atlantis using the submarine Alvin (cruise AT11-07, dive 3968) in 2004. DNA was extracted from basalt chips using a phenol-chloroform extraction including a negative control. DNA was amplified using the illustrate GenomiPhi V2 DNA Multiple Displacement Amplification (MDA) kit (GE Healthcare Life Sciences, Pittsburgh, PA, USA). Final DNA samples and the control were sent to the core genomics center at University of Pennsylvania for whole genome shotgun sequencing on a Roche GS-FLX Titanium 454 sequencer (454 Life Sciences, Branford, CT, USA).
Loi’hi: Two seafloor basalt samples (J2-243 R2-F, J-246 R2) were collected from the Lō’ihi Seamount (18.47 N, 155.18 W) at a depth of 5,000 m aboard the R/V Melville using the ROV Jason II in 2006. DNA was amplified using the illustrate GenomiPhi V2 DNA Multiple Displacement Amplification (MDA) kit (GE Healthcare Life Sciences, Pittsburgh, PA, USA). Final DNA samples and the control were sent to the core genomics center at University of Pennsylvania for whole genome shotgun sequencing on a Roche GS-FLX Titanium 454 sequencer (454 Life Sciences, Branford, CT, USA).
Negative control: DNA was extracted from sterile MilliQ water using a phenol-chloroform extraction. DNA was amplified using the illustrate GenomiPhi V2 DNA Multiple Displacement Amplification (MDA) kit (GE Healthcare Life Sciences, Pittsburgh, PA, USA). Final DNA was sent to the core genomics center at University of Pennsylvania for whole genome shotgun sequencing on a Roche GS-FLX Titanium 454 sequencer (454 Life Sciences, Branford, CT, USA).
Processing Description
All seafloor basalts were stored frozen at -80°C for XRD analysis and DNA extraction. Bulk mineralogy analysis, i.e. quantitative determination of rock-forming minerals and total clay minerals, was determined on all three seafloor basalts via X-ray Diffraction (XRD) analysis at KT GeoServices, Inc. Detection limits were at 1-5 wt%. For the two Lō’ihi seafloor basalts, average values were used.
Raw sequence reads were evaluated with FastQC version 0.11.3 (Schmieder and Edwards, 2011a), quality trimmed (minimum quality score – 25, maximum length – 450 bp, maximum homopolymer length – 9 bp, max N-tail – 1 bp), and filtered (removal of technical duplicates, minimum length – 60 bp) with Prinseq 0.20.4 (Schmieder and Edwards, 2011b) and MG-RAST (Meyer et al., 2008). We obtained 1,191,651 sequences in the EPR dataset; 1,102,191 sequences in the Lo’ihi dataset; and 58,188 sequences in the negative control dataset. Quality-filtered reads were assembled denovo using standard 454 settings in mira 3.4.1.1 (Chevreux et al., 1999). Padded (i.e. including potential gaps) contigs > 500 bp were filtered using mira 3.4.1.1 (convert_project) (Chevreux et al., 1999). Seafloor basalt contigs were screened for contamination using a combination of BBMap (bbduk.sh with parameters mcf=0.25, k=31) and the BLASTN algorithm (Altschul et al., 1990). The BBMap algorithm identified 4 potentially contaminant contigs in the EPR metagenome dataset (total of 4,290 bp) and 10 potentially contaminant contigs in the Lo’ihi (total of 12,423 bp). Community richness was estimated using the Chao1 index, diversity analysis was calculated using the Shannon index in QIIME 1.9.1 (alpha_diversity.py) based on BLASTX assignments of contigs. Phylosift was used to assess community diversity using the core molecular marker set of genes, which includes ~40 three-domain protein coding genes, single-copy eukaryote specific nuclear orthologs, ribosomal RNA genes (16S/18S), mitochondrial genes (mtDNA markers), and plastid and viral markers identified through Markov-clustering algorithms applied to genome datasets (Darling et al., 2014).
BCO-DMO Processing:
version 2015-11-06: replaced version 2015-10-23. Added site, lat, lon, and date columns.
Instruments
DNA was amplified using the illustrate GenomiPhi V2 DNA Multiple Displacement Amplification (MDA) kit (GE Healthcare Life Sciences, Pittsburgh, PA, USA)
General term for a laboratory apparatus commonly used for performing polymerase chain reaction (PCR). The device has a thermal block with holes where tubes with the PCR reaction mixtures can be inserted. The cycler then raises and lowers the temperature of the block in discrete, pre-programmed steps.
(adapted from http://serc.carleton.edu/microbelife/research_methods/genomics/pcr.html)
Roche GS-FLX Titanium 454 sequencer (454 Life Sciences, Branford, CT, USA) at the core genomics center at University of Pennsylvania.
General term for a laboratory instrument used for deciphering the order of bases in a strand of DNA. Sanger sequencers detect fluorescence from different dyes that are used to identify the A, C, G, and T extension reactions. Contemporary or Pyrosequencer methods are based on detecting the activity of DNA polymerase (a DNA synthesizing enzyme) with another chemoluminescent enzyme. Essentially, the method allows sequencing of a single strand of DNA by synthesizing the complementary strand along it, one base pair at a time, and detecting which base was actually added at each step.
X-ray Diffraction (XRD) analysis at KT GeoServices, Inc.
Instruments that identify crystalline solids by measuring the characteristic spaces between layers of atoms or molecules in a crystal.
Parameters
Site name: EPR = East Pacific Rise: Loihi = Loi'hi Seamount Hawaii.
latitude, in decimal degrees, North is positive, negative denotes South; Reported in some datasets as degrees, minutes
longitude, in decimal degrees, East is positive, negative denotes West; Reported in some datsets as degrees, minutes
Link to file (such as a data file or document).
link to NCBI GenBank accession page
Dataset Maintainers
Name | Affiliation | Contact |
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Katrina J. Edwards | University of Southern California (USC) | |
Esther Singer | University of Southern California (USC) | |
Nancy Copley | University of Southern California (USC) | ✓ |
BCO-DMO Project Info
Project Title | Metagenomic signatures in seafloor rocks and subsurface sediments: East Pacific Rise and Loihi Seamount |
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Acronym | EPR and Loihi basalt genomes |
URL | https://www.bco-dmo.org/project/616350 |
Created | October 23, 2015 |
Modified | November 6, 2015 |
Project Description
The seafloor and subsurface microbial world represents a significant portion of life on our planet. The influence on its proximate ambience and global processes, such as element cycles, has potentially been largely underestimated and not always been precisely evaluated. I am interested in the nature of deep biosphere microorganisms in rocks from the Loihi seamount, Hawai’i, the East Pacific Rise, and the Juan de Fuca Ridge, as well as in sediments from North Pond (Mid-Atlantic). In order to assess microbial diversity, metabolic activity, adaptation strategies and biogeographical signatures in the deep subseafloor biosphere, metagenomics by pyrosequencing will be used to complement previous research efforts with the most in-depth and precise data that is available to date.
This project is a C-DEBI Graduate Student Fellowship award (2011)
Data Project Maintainers
Name | Affiliation | Role |
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Esther Singer | University of Southern California (USC) | Principal Investigator |