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Palmer LTER 2014 cruise microbial Eukaryote (18s) plankton
Citation
Lin Y, Cassar N, Marchetti A, Moreno C, Ducklow H, Li Z (2018): Palmer LTER 2014 cruise microbial Eukaryote (18s) plankton. v1.3. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=palmer_lter_eukaryote_18s_plankton&v=1.3 https://doi.org/10.15468/wtt5gg
Contact: Yajuan, Lin

Access data
Archived data
Availability: Creative Commons License This dataset is licensed under a Creative Commons Attribution 4.0 International License.

Description
Amplicon sequencing dataset of microbial eukaryotes (18S ssh rRNA v4)living in the plankton of the Southern Ocean around the Western Antarctic Peninsula. Samples were collected during the annual Palmer LTER cruise from Jan to early Feb in 2014 aboard the R/V Laurence M. Gould. more

Surface seawater from the vessel seawater supply line was gently vacuum filtered through a 0.2 µm pore size Millipore Supor filters. Each filter was then preserved immediately in 1 ml of RNA-later and stored at −80 °C.
Samples were taken during the annual Palmer LTER cruise from Jan to early Feb in 2014 aboard the R/V Laurence M. Gould.
Method step description:
  1. From each sampled station one filter was first split in two halves, one for DNA extraction and the other for later RNA studies. 500 µl of the RNAlater from the original tube was loaded onto an Amicon 10 k column and the column was spun for 15 minutes at 14,000 g to get rid of most of the RNAlater and concentrate cells suspended in RNAlater. The resulting solution (~50 µl) was then added back to the one half filter. The cells on filter were lysed by bead-beating using 0.2 g of zirconium beads in 400 µl of buffer AP1 (Qiagen).
  2. DNA was extracted and purified using Qiagen DNeasy Plant Mini kit following the manufacture’s instruction. 18 S (eukaryotic) rDNA gene fragments were amplified by PCR using V4 primer sets: 18 S forward (5′- CCAGCASCYGCGGTAATTCC-3′) and reverse (5′-ACTTTCGTTCTTGAT-3′). Amplicons from each sample were tagged using dual index fusion primers and a “heterogeneity spacer” was used to improve sequencing quality. The “heterogeneity spacer” is a 0–5 bp of spacer added to the index sequence in order to allow different samples be sequenced out of phase and thus improve the amplicon library sequencing quality. Each PCR reaction (25 µl) consisted of 1U of Platinum Taq DNA Polymerase High Fidelity (Invitrogen), 1 × High Fidelity Buffer, 200 µM dNTPs, 2 mM MgSO4, 0.2 µM each primer, and 5–30 ng extracted environmental DNA template. PCR was conducted with an initial activation step at 94 °C for 3 min, followed by 30 three-step cycles consisting of 94 °C for 30 s, 57 °C for 30 s and 72 °C for 1 min, and a final extension step of 72 °C at 10 min. PCR products in triplicates for each sample were then pooled and purified using QIAquick PCR Purification kit, and the concentration of amplicon DNA was quantified using a Qubit dsDNA assay.
  3. Amplicons from different locations were pooled in equimolar amounts (final concentration ~10 ng/µl) and submitted to Duke Institute for Genomic Sciences and Policy (IGSP) for sequencing using Illumina MiSeq. 300PE platform.

Scope
Themes:
Biology > Plankton
Keywords:
Marine/Coastal, Eukaryotes, Metadata, PS, Southern Ocean

Geographical coverage
PS, Southern Ocean [Marine Regions]

Temporal coverage
January 2014 - February 2014

Parameter
Molecular data

Contributors
Duke University, moredata creator

Related datasets
Published in:
AntOBIS: Antarctic Ocean Biodiversity Information System, more
(Partly) included in:
RAS: Register of Antarctic Species, more

Dataset status: Completed
Data type: Metadata
Data origin: Research: field survey
Metadatarecord created: 2018-12-12
Information last updated: 2019-04-10
All data in the Integrated Marine Information System (IMIS) is subject to the VLIZ privacy policy