元數據

Bacteria (16S ssu rRNA) in an Antarctic snow sample

最新版本 由 SCAR - Microbial Antarctic Resource System 發佈於 2019年3月19日 SCAR - Microbial Antarctic Resource System
發布日期:
2019年3月19日
授權條款:
CC-BY 4.0

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說明

Amplicon sequencing sample of Bacteria (16S ssu rRNA gene, v1-v3 region) from a snow sample taken from the "clean Area”, 2 km South from the Antarctic Research Base “Concordia” (75°06′S–123°20′E).

版本

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如何引用

研究者應依照以下指示引用此資源。:

Michaud L, Lo Giudice A, Mysara M, Monsieurs P, Raffa C, Leys N, Almafitano S, Van Houdt R (2019): Bacteria (16S ssu rRNA) in an Antarctic snow sample. v1.2. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=bacteria_antarctic_snow&v=1.2

權利

研究者應尊重以下權利聲明。:

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: 647cb838-3294-4ee3-8ebc-a3a0075c7e5a。  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

資源建立者:
-

Luigi Michaud
University of Messina
Messina
IT
Angelina Lo Giudice
University of Messina
Messina
IT
Mohamed Mysara
Vrije Universiteit Brussel
Brussels
BE
Pieter Monsieurs
Belgian Nuclear Research Centre (SCK CEN)
Mol
BE
Carmella Raffa
University of Messina
Messina
IT
Natalie Leys
Belgian Nuclear Research Centre (SCK CEN)
Mol
BE
Stefano Almafitano
National Research Council (IRSA-CNR)
Rome
IT
Rob Van Houdt
Belgian Nuclear Research Centre (SCK CEN)
Mol
BE

可回覆此資源相關問題者:

Luigi Michaud
University of Messina
Messina
IT
Angelina Lo Giudice
University of Messina
Messina
IT

元數據填寫者:
-

Maxime Sweetlove
Research assistent
Royal Belgian Institute for Natural Sciences
Rue Vautier 29
Brussels

與此資源的相關者:

使用者

地理涵蓋範圍

Antarctic snow surface sample collected in the “Clean Area” 2 km South from the Research Base “Concordia” (75°06′S–123°20′E)

界定座標範圍 緯度南界 經度西界 [-75, -123], 緯度北界 經度東界 [-75, -123]

分類群涵蓋範圍

Bacteria (16S ssu rRNA gene, v1-v3 region)

Domain Bacteria (Bacteria)

時間涵蓋範圍

起始日期 2010-01-01

計畫資料

無相關描述

計畫名稱 Bacteria (16S ssu rRNA) in an Antarctic snow sample
經費來源 Support was given by the Italian “Programma Nazionale di Richerche in Antartide” (PNRA) and the MNA (Museo Nazionale dell'Antartide)

參與計畫的人員:

Luigi Michaud
Angelina Lo Giudice

取樣方法

Sampling was performed by using polyethylene boxes pre-treated with 1M hydrogen chloride and hydrogen peroxide. Sterile gloves and suit, and an ethanol flame-sterilized shovel were used.

研究範圍 Snow surface sample was collected in triplicate from a “Clean Area” 2 km from the Research Base “Concordia” (75°06′S–123°20′E)
品質控管 Autoclave-sterilized Milli-Q water was treated in tandem with the snow samples as a negative-control field blank. Quantity and quality of extracted DNA was checked by nanodrop ND-1000 device and the Quant-iT PicoGreen dsDNA reagent and kit (Life Tech, Carlsbad, USA) following the manufacturer's instructions.

方法步驟描述:

  1. Collected samples were allowed to thaw at 4°C for 24–48 h in the laboratory, with 100 litres of packed snow per sample resulting in approximately 20 litres of snowmelt.
  2. 15 L melted snow for DNA extraction was filtered through a 0.2-µm-pore-size Sterivex filter unit (Millipore). The filters were stored at −20°C in lysis buffer (50 mM tris, 40 mM EDTA, and 750 mM sucrose).
  3. Genomic DNA was extracted in triplicate using the phenol-chloroform method according to Zhou et al., and precipitated by adding 0.7 volumes of 100% isopropanol followed by a wash with ice-cold 70% ethanol. After air-drying, DNA was resuspended in 50 µl of deionizated sterile water.
  4. PCR of a bacterial 16S rRNA gene fragment (V1–V3 region, 507 bp) and subsequent tag-encoded pyrosequencing were performed at DNAVision (Charleroi, Belgium). The 16S rRNA genes were amplified using the two universal primers 8F (5′- AGAGTTTGATCCTGGCTCAG -3′) and 518R (5′- ATTACCGCGGCTGCTGG -3′). The forward primer contained the sequence of the Titanium A adaptor (5′-CCATCTCATCCCTGCGTGTCTCCGACTCAG-3′) and a barcode sequence. For each sample, a PCR mix of 100 µl was prepared containing 1×PCR buffer, 2U of KAPA HiFi Hotstart polymerase blend and dNTPs (Kapabiosystems), 300 nM primers (Eurogentec, Liege, Belgium), and 60 ng gDNA. Thermal cycling consisted of initial denaturation at 95°C for 5 min, followed by 25 cycles of denaturation at 98°C for 20 s, annealing at 56°C for 40 s, and extension at 72°C for 20 s, with a final extension of 5 min at 72°C. 3 µl of PCR product were added to a new PCR mix (identical as first round of PCR) for the nested PCR of 15 cycles. Amplicons were visualized on 1% agarose gels using GelGreen Nucleic Acid gel stain in 1× TAE (Biotium) and were cleaned using the Wizard SV Gel and PCR Clean-up System (Promega) according to the manufacturer's instructions. Pyrosequencing was carried out using the forward primer on a 454 Life Sciences Genome Sequencer FLX instrument (Roche) following titanium chemistry.

引用文獻

  1. Michaud, L., Giudice, A. L., Mysara, M., Monsieurs, P., Raffa, C., Leys, N., ... & Van Houdt, R. (2014). Snow surface microbiome on the High Antarctic Plateau (DOME C). PloS one, 9(8), e104505.

額外的詮釋資料

替代的識別碼 647cb838-3294-4ee3-8ebc-a3a0075c7e5a
https://ipt.biodiversity.aq/resource?r=bacteria_antarctic_snow