URLhttps://www.bco-dmo.org/dataset/616326
Download URLhttps://www.bco-dmo.org/dataset/616326/data/download
Media Typetext/tab-separated-values
CreatedOctober 23, 2015
ModifiedAugust 19, 2016
StateFinal no updates expected
Brief DescriptionAssembled 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

Jason II [ROV Jason]
Details
Instance Description (Jason II)

Jason dives 243 and 246 (18.47 N, 155.18 W).

The Remotely Operated Vehicle (ROV) Jason is operated by the Deep Submergence Laboratory (DSL) at Woods Hole Oceanographic Institution (WHOI). WHOI engineers and scientists designed and built the ROV Jason to give scientists access to the seafloor that didn't require them leaving the deck of the ship. Jason is a two-body ROV system. A 10-kilometer (6-mile) fiber-optic cable delivers electrical power and commands from the ship through Medea and down to Jason, which then returns data and live video imagery. Medea serves as a shock absorber, buffering Jason from the movements of the ship, while providing lighting and a bird’s eye view of the ROV during seafloor operations. During each dive (deployment of the ROV), Jason pilots and scientists work from a control room on the ship to monitor Jason’s instruments and video while maneuvering the vehicle and optionally performing a variety of sampling activities. Jason is equipped with sonar imagers, water samplers, video and still cameras, and lighting gear. Jason’s manipulator arms collect samples of rock, sediment, or marine life and place them in the vehicle’s basket or on "elevator" platforms that float heavier loads to the surface. More information is available from the operator site at URL.
Instance Description

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)

Instance Description

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 diffractometer (XRD) [X-ray diffractometer]
Details
Instance Description (X-ray diffractometer (XRD))

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 [site]
Details
site

Site name: EPR = East Pacific Rise: Loihi = Loi'hi Seamount Hawaii.  

Sampling site identification.
lat [latitude]
Details
lat
latitude; north is positive

latitude, in decimal degrees, North is positive, negative denotes South; Reported in some datasets as degrees, minutes

lon [longitude]
Details
lon
longitude;east is positive

longitude, in decimal degrees, East is positive, negative denotes West; Reported in some datsets as degrees, minutes

year [year]
Details
year
year of sample collection

year, reported as YYYY, e.g. 1995

month [month]
Details
month
month of sample collection

Month of year; numeric 1 to 12.

day [day]
Details
day
day of sample collection
day of month; numeric 1 to 31
sample_type [sample_type]
Details
sample_type
type of sample
text description of type of sample collected (rock, bio, fluid, etc)
Fasta_file_link [file_link]
Details
Fasta_file_link
Link to FASTA metagenome files

Link to file (such as a data file or document).

NCBI_accession [accession number]
Details
NCBI_accession

link to NCBI GenBank accession page

Database identifier assigned by repository and linked to GenBank or other repository.

Dataset Maintainers

NameAffiliationContact
Katrina J. EdwardsUniversity of Southern California (USC)
Esther SingerUniversity of Southern California (USC)
Esther SingerUniversity of Southern California (USC)
Nancy CopleyWoods Hole Oceanographic Institution (WHOI BCO-DMO)

BCO-DMO Project Info

Project TitleMetagenomic signatures in seafloor rocks and subsurface sediments: East Pacific Rise and Loihi Seamount
AcronymEPR and Loihi basalt genomes
URLhttps://www.bco-dmo.org/project/616350
CreatedOctober 23, 2015
ModifiedNovember 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)

Project Maintainers
NameAffiliationRoleContact
Esther SingerUniversity of Southern California (USC)Principal Investigator
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