Dataset: Assembled metagenome sequences > 500 bp
Deployment: AT11-07

Assembled metagenome sequences > 500 bp from EPR, Lo'ihi, and control

Lead Principal Investigator:

Katrina Edwards (University of Southern California, USC)

Principal Investigator:

Esther Singer (University of Southern California, USC)

Contact:

Esther Singer (University of Southern California, USC)

BCO-DMO Data Manager:

Nancy Copley (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Project:

Metagenomic signatures in seafloor rocks and subsurface sediments: East Pacific Rise and Loihi Seamount (EPR and Loihi basalt genomes)

Current State:

Final no updates expected

Version:

2015-10-23

Coordinated Deployments:

AT11-07_Alvin_Dives

Version Date:

2015-10-23

Data URL:

https://www.bco-dmo.org/dataset-deployment/616336/data

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Description

This dataset includes FASTA formatted files of the metagenomes from the East Pacific Rise seafloor basalt, the Loi'hi Seamount seafloor basalt, and negative control of sterile water.

The sequence data has been submitted to NCBI under Bioproject number PRJNA302671:
- Biosamples SUB1190178 (blank)
- SUB1190153 (Lo'ihi)
- SUB1190159 (EPR)

This dataset is the result of a C-DEBI Graduate Student Fellowship award (2011).

Methods & Sampling

Dataset 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).

Data Processing Description

Dataset 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.

More information about this dataset deployment

Funding

Award Number	Funding Source
OCE-0939564	NSF Division of Ocean Sciences

Instruments

Automated DNA Sequencer

Supplied Name:

Supplied Description:

Roche GS-FLX Titanium 454 sequencer (454 Life Sciences, Branford, CT, USA) at the core genomics center at University of Pennsylvania.

Instrument Type

Generic Name: Automated DNA Sequencer

Acronym: Automated Sequencer

Generic Description:

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.

ROV Jason

Supplied Name: Jason II

Supplied Description:

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

Instrument Type

Generic Name: ROV Jason

Generic Description:

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. https://ndsf.whoi.edu/jason/

Thermal Cycler

Supplied Name:

Supplied Description:

DNA was amplified using the illustrate GenomiPhi V2 DNA Multiple Displacement Amplification (MDA) kit (GE Healthcare Life Sciences, Pittsburgh, PA, USA)

Instrument Type

Generic Name: Thermal Cycler

Generic Description:

A thermal cycler or "thermocycler" is a general term for a type of laboratory apparatus, commonly used for performing polymerase chain reaction (PCR), that is capable of repeatedly altering and maintaining specific temperatures for defined periods of time. 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. They can also be used to facilitate other temperature-sensitive reactions, including restriction enzyme digestion or rapid diagnostics.

(adapted from http://serc.carleton.edu/microbelife/research_methods/genomics/pcr.html)

X-ray diffractometer

Supplied Name: X-ray diffractometer (XRD)

Supplied Description:

X-ray Diffraction (XRD) analysis at KT GeoServices, Inc.

Instrument Type

Generic Name: X-ray diffractometer

Acronym: XRD

Generic Description:

Instruments that identify crystalline solids by measuring the characteristic spaces between layers of atoms or molecules in a crystal.

Parameters

Supplied Name	Supplied description	Supplied Units	Standard Name
site	Site name: EPR = East Pacific Rise: Loihi = Loi'hi Seamount Hawaii.	unitless	site
lat	latitude; north is positive	decimal degrees	lat
lon	longitude;east is positive	decimal degrees	lon
year	year of sample collection	YYYY	year
month	month of sample collection	MM	month
day	day of sample collection	DD	day
sample_type	type of sample	unitless	sample_type
Fasta_file_link	Link to FASTA metagenome files	unitless	file_link
NCBI_accession	link to NCBI GenBank accession page	unitless	accession_number

Database

Contribute Data

Dataset: Assembled metagenome sequences > 500 bp
Deployment: AT11-07

Dataset acquisition description

Dataset Processing Description

Database

Contribute Data

Dataset: Assembled metagenome sequences > 500 bpDeployment: AT11-07

Dataset acquisition description

Dataset Processing Description

Dataset: Assembled metagenome sequences > 500 bp
Deployment: AT11-07