AT26-18_FLOCS_DNA16S

URL	https://www.bco-dmo.org/dataset/754354
Created	January 29, 2019
Modified	February 20, 2019
State	Final no updates expected
Brief Description	16S rRNA gene amplicon sequences from FLOCS experiments at the Juan de Fuca Ridge CORKs

Acquisition Description

Polished mineral chips were prepared and deployed as described in Fisher et al. 2011 (DOI: 10.2204/iodp.proc.327.107.2011). Mineral colonization experiments were recovered during cruise AT26-18 in August 2014 and processed as described in Orcutt et al. 2011 (DOI: 10.1038/ismej.2010.157). Approximately 6g of each substrate underwent total DNA extraction using the MOBIO PowerSoil® DNA Isolation Kit (MOBIO Laboratories, Inc.) with modification from manufacturer instructions to include a phenol:chloroform:isoamyl alcohol (24:25:1) extraction step. Each batch of samples included procedural blanks. Replicate extracts were pooled and concentrated to 100 μl with vacuum centrifugation (SavantTM DNA SpeedVacTM Concentrator). Concentrated extracts were cleaned with the CleanAll DNA/RNA Clean Up and Concentration Kit (Norgen Biotek Corp, Thorold, Canada) and eluted in 20-50 μl of PCR-grade water. Sequencing of the V1-V3 region of the 16S rRNA gene was performed by Research and Testing Laboratory (Lubbock, TX, USA) using the forward primer 28F (5′- GAG TTT GAT CNT GGC TCA G -3′) and reverse primer 388R (5’- TGC TGC CTC CCG TAG GAG T -3’) for Bacteria using 2 x 300 bp kits on the Illumina MiSeq platform.

Processing Description

BCO-DMO Processing Notes:
– Nothing to report

Instruments

Illumina MiSeq [Automated DNA Sequencer]

Details

Instance Description (Illumina MiSeq)

Concentrated extracts were cleaned with the CleanAll DNA/RNA Clean Up and Concentration Kit (Norgen Biotek Corp, Thorold, Canada) and eluted in 20-50 μl of PCR-grade water. Sequencing of the V1-V3 region of the 16S rRNA gene was performed by Research and Testing Laboratory (Lubbock, TX, USA) using the forward primer 28F (5′- GAG TTT GAT CNT GGC TCA G -3′) and reverse primer 388R (5’- TGC TGC CTC CCG TAG GAG T -3’) for Bacteria using 2 x 300 bp kits on the Illumina MiSeq platform.

Automated DNA Sequencer

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.

Parameters

Dataset Maintainers

Name	Affiliation	Contact
Beth N. Orcutt	Bigelow Laboratory for Ocean Sciences	✓
Gustavo A. Ramírez	Bigelow Laboratory for Ocean Sciences	✓
Mathew Biddle	University of Southern California (USC)
Mathew Biddle	University of Southern California (USC)
Mathew Biddle	Woods Hole Oceanographic Institution (WHOI BCO-DMO)

BCO-DMO Project Info

Project Title	Collaborative Research: Completing single- and cross-hole hydrgeologic and microbial experiments: Juan de Fuca Flank
Acronym	JdF Flank
URL	https://www.bco-dmo.org/project/625989
Created	October 29, 2015
Modified	December 21, 2018

Project Description

NSF Award Abstract:
In 2010, IODP exp. 327 launched series of experiments based on instrumentation placed in 6 boreholes (Holes 1026B, 1027C, 1301A, 1301B, 1362A, and 1362B) on and near the Juan de Fuca Ridge (JDFR). The experiments were designed to examine hydrological and biological processes in basaltic ocean crust. Major issues to be addressed include overall permeability, the magnitudes, velocity and directions of fluid, solute,and heat transport,constraints on the fluid storage properties of crust, determining how fluid reservoirs respond to seismic perturbations, and investigating the variability, diversity, and metabolism of resident microbial populations. This project will recover sensors and samplers in the instrumented boreholes(CORKs),access data and samples, collect biological incubators placed at depth within the boreholes, and seal the CORKs. All of these activities are needed for the completion of the JDFR experiments

Once the data and samples are retrieved, the PI’s will develop new numerical models for the hydrology (permeability, thickness and extent of permeable zones, connectivity, direction of flow, and anisotropy) of ocean crust based on the results of flow tests. T sensors will enable thermal structure to be determined in conjunction with flow parameters. DNA will be analyzed via 16S ribosomal RNA (rRNA) genes as well as select full-length Sanger-style sequencing and gene fingerprinting techniques. The PI’s will look at diversity, shared species, emergence and disappearance of groups. Metagenomic work will be done on some samples.

Broader Impacts include training of 4 shipboard educators, with an emphasis on recruiting underrepresented groups for these positions, teacher workshops, and special training for undergraduate and graduate students and post-docs. Research team includes grads, undergrads, and post-docs

Data Project Maintainers

Name	Affiliation	Role
Charles Geoffrey Wheat	University of Alaska Fairbanks (UAF-IMS)	Lead Principal Investigator
Katrina J. Edwards	University of California-Santa Barbara (UCSB)	Principal Investigator
Andrew T. Fisher	University of Miami Rosenstiel School of Marine and Atmospheric Science (UM-RSMAS)	Principal Investigator
Jordan F. Clark	University of Southern California (USC)	Principal Investigator
Keir Becker	University of California-Santa Cruz (UC Santa Cruz)	Principal Investigator
Michael S. Rappé	University of Hawaii at Manoa (HIMB)	Co-Principal Investigator

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