Palmer LTER 2014 cruise microbial Eukaryote (18s) plankton
Citation
Lin Y, Cassar N, Marchetti A, Moreno C, Ducklow H, Li Z, Sweetlove M (2019). Palmer LTER 2014 cruise microbial Eukaryote (18s) plankton. Version 1.3. SCAR - Microbial Antarctic Resource System. Metadata dataset https://doi.org/10.15468/wtt5gg accessed via GBIF.org on 2024-08-31.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.Sampling Description
Study Extent
Samples were taken during the annual Palmer LTER cruise from Jan to early Feb in 2014 aboard the R/V Laurence M. Gould.Sampling
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.Method steps
- 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).
- 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.
- 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.
Taxonomic Coverages
microbial eukaryote plankton (18S ssh rRNA gene v4 region)
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Eukaryotacommon name: Eukaryotes rank: domain
Geographic Coverages
Southern Ocean around the Western Antarctic Peninsula.
Bibliographic Citations
- Lin, Y., Cassar, N., Marchetti, A., Moreno, C., Ducklow, H., & Li, Z. (2017). Specific eukaryotic plankton are good predictors of net community production in the Western Antarctic Peninsula. Scientific reports, 7(1), 14845. -
Contacts
Yajuan Linoriginator
Duke University
Durham
US
Nicolas Cassar
originator
Duke University
Durham
US
Adrian Marchetti
originator
University of North Carolina at Chapel Hill
Chapel Hill
US
Carly Moreno
originator
University of North Carolina at Chapel Hill
Chapel Hill
US
Hugh Ducklow
originator
Columbia University
Palisades
US
Zuchuan Li
originator
Duke University
Durham
US
Maxime Sweetlove
metadata author
position: Research assistent
Royal Belgian Institute for Natural Sciences
Rue Vautier 29
Brussels
1000
BE
email: msweetlove@naturalsciences.be
Yajuan Lin
administrative point of contact
Duke University
Durham
US