Arctic multi-year sea ice Bacteria
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Hatam I, Lange B, Beckers J, Haas C, Lanoil B, Charchuk R (2019): Arctic multi-year sea ice Bacteria. v1.2. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=arctic_multiyear_sea_ice_bacteria&v=1.2
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Landfast ice off the northern shore of northern Ellesmere Island Nunavut, Canada (82.54905°N, −62.37685°W).
|Bounding Coordinates||South West [82.549, -62.377], North East [82.549, -62.377]|
Bacteria, targeted with the 16S ssu rRNA marker gene (v1-v3 region)
No Description available
|Title||Arctic multi-year sea ice Bacteria|
|Funding||Support was provided by the Natural Science and Engineering Research Council of Canada (NSERC) Discovery Grant program and the Polar Continental Shelf Program (PCSP); the Northern Scientific Training Program (NSTP) and Circumpolar-Boreal Arctic Research (C-BAR) programs.|
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Two parallel cores were sampled using Kovacs Mark II 9 cm corer (Kovacs Enterprise, Roseburg, Oregon) and a 36 V electric hand drill. Prior to drilling, the core barrel was rinsed with sterile deionized water (Milli-Q Integral Water Purification System, EMD Millipore Corporation, Billerica, MA). The two cores were immediately sectioned on site to 30-cm intervals using a hand saw (rinsed in deionized water and wiped with an ethanol wipe) and placed in UV-sterilized polypropylene bags.
|Study Extent||Two ice cores were taken during May 2011 at one site on landfast ice off the northern shore of northern Ellesmere Island Nunavut, Canada (82.54905°N, −62.37685°W).|
Method step description:
- Ice samples used were thawed at room temperature in the dark, and were filtered individually through 0.22-μm-pore-size polyethersulfone membrane filters (Pall, Mississauga, ON, Canada). Direct melting was used to avoid nonspecific addition of DNA and dilution of samples. All glassware was sterilized using 10% household bleach solution followed by thorough washing in sterile deionized water between samples to prevent cross-contamination. Each filter was placed in a microfuge tube and submerged in RNAlater® solution (Life Technologies, Burlington, ON, Canada) and stored at −20 °C for later DNA extraction.
- DNA was extracted from preserved filters by bead beating using the FastDNA® SPIN Kit for Soil (MP Biomedicals, Solon, OH) as instructed by the manufacturer. Filters from equivalent 30-cm sections of duplicate ice cores were combined prior to DNA extraction to ensure sufficient DNA yield. Seawater duplicate samples were processed individually.
- Molecular Research LP (Shallowater, TX) performed pyrosequencing of the V1-V3 regions of the bacterial 16S rRNA gene as described in Dowd et al. (2008). Amplification was performed using HotStarTaq Plus Master Mix Kit (Qiagen, Valencia, CA) under the following conditions: 94 °C for 3 min, followed by 28 cycles of 94 °C for 30 s; 53 °C for 40 s; and 72 °C for 1 min with a final elongation step at 72 °C for 5 min. Gene-specific forward PCR primer sequences were tagged with the sequencing adapters for GS FLX Titanium chemistry, an 8 base barcode, and a linker sequence. All PCRs were performed in triplicate, mixed in equal concentrations, and purified using Agencourt AMPure beads (Agencourt Bioscience Corporation, MA). FLX-Titanium amplicon pyrosequencing was performed using the Genome Sequencer FLX System (Roche, Branford, CT).
- Hatam, I., Charchuk, R., Lange, B., Beckers, J., Haas, C., & Lanoil, B. (2014). Distinct bacterial assemblages reside at different depths in Arctic multiyear sea ice. FEMS microbiology ecology, 90(1), 115-125. https://doi.org/10.1111/1574-6941.12377