algal_mat_microbiome_east_antarctica

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如何引用

研究者應依照以下指示引用此資源。:

Hirose Y, Shiozaki T, Otani M, Kudoh S, Imura S, Eki T, Harada N (2021): algal_mat_microbiome_east_antarctica. v1.0. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=algal_mat_microbiome_east_antarctica&v=1.0

權利

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此資料的發布者及權利單位為 SCAR - Microbial Antarctic Resource System。 This work is licensed under a Creative Commons Attribution (CC-BY 4.0) License.

GBIF 註冊

此資源已向GBIF註冊，並指定以下之GBIF UUID: 5d7ab0cf-4c60-4b8d-bea4-04632af68fc9。  SCAR - Microbial Antarctic Resource System 發佈此資源，並經由Scientific Committee on Antarctic Research同意向GBIF註冊成為資料發佈者。

關鍵字

Metadata

聯絡資訊

地理涵蓋範圍

East Antarctica, Langhovde and Skarvnes region

時間涵蓋範圍

計畫資料

無相關描述

參與計畫的人員:

取樣方法

Microbial mat samples were collected from 1–2 cm of the surface using a scoop. Water samples were collected using a disposable plastic syringe. Samples were transferred to the icebreaker SHIRASE and then frozen until DNA extraction was performed. Sample S1 was a brown mat isolated from Lake Mitsu Ike; S2 was a red colored bloom observed in a puddle of thawing snow in the coastal area; S3 was a green and brown mat that was collected in Lake Yukidori Ike; S4 was a black microbial mat in Lake Bosatsu Ike; S5 was a small white filamentous aggregate floating on the surface of Lake Bosatsu Ike; S6 was a brown microbial mat in Lake Bosatsu Ike; S7 was a green colored aggregate found in a small stream near Lake Suribati Ike; S8 was a brown mat found near the stream; S9 was a white aggregate floating on the surface of Lake Suribati Ike; S10 was a black and brown mat collected from Lake Neko Ike; S11 was a floating brown mat that originated from the benthic algal mat in Lake Kobachi Ike; S12 was a dark yellow mat collected from thawed soil near Lake Tokkuri Ike; and S13 was a orange mat, found at the bottom of the shallows in Lake Kumogata Ike.

方法步驟描述:

1. The samples were mixed with 10 mL of 10 mM Tris-HCl pH 8.0, 5 mL of phenol pH 8.0, and 2 g of zirconia/glass beads (diameter 0.1 mm), and vortexed vigorously for three minutes at room temperature. After heat treatment at 65 ◦C for 10 min, the debris and beads were removed by centrifugation for five minutes at 16,000× g. The upper water phase was transferred to a new tube and an equal volume of chloroform/isoamylalchol (24:1) was added, vortexed vigorously, and centrifuged for five minutes at 16,000× g. The upper water phase was precipitated with 2.5 volumes of 99.5% ethanol and 0.1 volumes of sodium acetate pH 5.2, and precipitated with centrifugation for 10 min at 21,600× g. The white pellets containing genomic DNA were washed with 70% ethanol, dried for five minutes at room temperature, and dissolved with 300 μl of 10 mM Tris-HCl pH 8.5. DNA was further purified using a DNeasy Blood and Tissue kit (Qiagen, Hilden, Germany), in accordance with the manufacturer’s instructions, and eluted with water.
2. The V3–V4 region of the 16S rRNA and the V7–V8 region of the 18 rRNA genes were amplified using KOD FX Neo (Toyobo, Osaka, Japan). Primer sets 341F and 805R were used for 16S rRNA analysis and F1183 and R1631 for 18S rRNA. PCR thermal cycle was an initial denaturing step at 94 ◦C for two minutes, 35 cycles of denaturation at 98 ◦C for 10 s, annealing at 55 ◦C for 30 s, and extension at 68 ◦C for 30 s, with the final extension step at 68 ◦C for five minutes. The PCR product was purified with 0.8 volumes of AMPure XP beads, in accordance with the manufacturer’s instructions, and eluted with 10 mM Tris-HCl pH 8.5. Index PCR was performed in eight cycles using a Nextera XT Index Kit v2 (Illumina, San Diego, California, USA), in accordance with the manufacturer’s instructions. The same index was used for the 16S and 18S rRNA amplicons that were obtained from the same sample. The amplified libraries were purified by the addition of 1.12 volumes of AMPure XP beads, in accordance with the manufacturer’s instructions, and eluted with 10 mM Tris-HCl pH 8.5. The concentration of each library was quantified using a spectrophotometer, and equal amounts of each library were pooled and quantified using a Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific, Waltham, Massachusetts, USA).
3. Each 300 bp end of the pooled library was sequenced using an MiSeq Reagent Kit v3 (600 cycles; Illumina) on the MiSeq instrument (Illumina).

引用文獻

1. Hirose, Y., Shiozaki, T., Otani, M., Kudoh, S., Imura, S., Eki, T., & Harada, N. (2020). Investigating algal communities in lacustrine and hydro-terrestrial environments of east Antarctica using deep amplicon sequencing. Microorganisms, 8(4), 497.

額外的詮釋資料