Aplicon sequencing dataset (454 pyrosequencing) of microbial Fungi (ITS) in soils from Bird Island, Signy Island and Leonie Island (Sub-Antarctica)
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Cox F, Newsham K, Robinson C (2019): Microbial Fungi in soils on different Sub-Antarctic islands. v1.0. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=microbial_fungi_from_3_sub_antarctic_islands&v=1.0
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Soils were sampled in Bird Island, Signy Island and Leonie Island (Sub-Antarctica)
|Bounding Coordinates||South West [-67.598, -68.356], North East [-54.009, -38.066]|
Fungi were profiled by sequencing the ITS gene (454 pyrosequencing)
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|Title||Microbial Fungi in soils on different Sub-Antarctic islands|
|Funding||Funding was provided by: the Antarctic Funding Initiative grant from the UK Natural Environment Research Council (grant numbers NE/H014098/1, NE/H014772/1 and NE/H01408X/1) and a British Ecological Society Large Research Grant for early career ecologists.|
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Soil was collected from under populations of co‐occurring Deschampsia antarctica Desv. and Colobanthus quitensis (Kunth) Bartl., the only two native vascular plant species that occur in Antarctica. On each island, 50 ml sterile centrifuge tubes (Corning Inc, Corning, NY, USA) were used to collect soil samples by hammering them directly into the vertical walls of three pits at three depths (2, 4 and 8 cm). The soil, kept on ice after collection and frozen at −80 °C within 5 h, was freeze dried to preserve fungal nucleotides.
|Study Extent||Soil samples were collected from Bird Island (54.0089°S, 38.0662°W), Signy Island (60.7107°S, 45.5849°W) and Léonie Island (67.5984°S, 68.3561°W) in the sub‐Antarctic, between October and November 2011.|
Method step description:
- Total DNA and RNA were extracted simultaneously from five individual 50 mg samples, taken from each of the tubes of homogenized soil (representing a total of 27 × 5 = 135 samples), using RNA PowerSoil Total RNA Isolation and DNA Elution Accessory kits (MoBio Laboratories, Carlsbad, CA, USA). Extracted DNA was amplified in triplicate PCR reactions using the primers ITS1F and ITS4 as described by Cox et al. (2016), with conditions matching those described below for cDNA. Extracted RNA was treated with a Turbo DNA‐free kit (Life technologies, Carlsbad, CA, USA), checked for the absence of DNA using PCR, and reverse transcribed using AccuScript High‐Fidelity Reverse Transcriptase (Agilent, Santa Clara, CA, USA) and random nonamers. The resulting cDNA was amplified in triplicate PCR reactions using ITS1F (Gardes and Bruns, 1993) and ITS4 (White et al., 1990) primers. The ITS4 primer was modified with the Roche 454 A adapter and a 10 bp barcode specific to each sample, allowing identification of different samples once pooled, and the ITS1F primer was modified with the 454 B adaptor.
- The triplicate PCR products were pooled and subsequently purified using AMPure XP bead purification (Beckman Coulter, Inc, Brea, CA, USA) and quantified using a Qubit dsDNA HS Assay (Life Technologies, Carlsbad, CA, USA) before normalization to consistent concentration. The purified and normalized PCR products were run on a single plate, on the 454 Roche Titanium FLX platform at the Liverpool Centre for Genomic Research, at the same time and under identical conditions to the DNA library.
- Cox, F., Newsham, K. K., & Robinson, C. H. (2019). Endemic and cosmopolitan fungal taxa exhibit differential abundances in total and active communities of Antarctic soils. Environmental microbiology. https://doi.org/10.1111/1462-2920.14533