Airborne Bacteria from Miers Valley, Antarctica

バージョン

次の表は、公にアクセス可能な公開バージョンのリソースのみ表示しています。

引用方法

研究者はこの研究内容を以下のように引用する必要があります。:

Bottos E, Woo A, Zawar-Reza P, Pointing S, Cary C (2018): Airborne Bacteria from Miers Valley, Antarctica. v1.2. SCAR - Microbial Antarctic Resource System. Dataset/Metadata. https://ipt.biodiversity.aq/resource?r=airborne_bacteria_miers_antarctica&v=1.2

権利

研究者は権利に関する下記ステートメントを尊重する必要があります。:

パブリッシャーとライセンス保持者権利者は SCAR - Microbial Antarctic Resource System。 This work is licensed under a Creative Commons Attribution (CC-BY 4.0) License.

GBIF登録

このリソースをはGBIF と登録されており GBIF UUID: 6a1177d0-ce25-4611-9666-a3d58cdea8c5が割り当てられています。   Scientific Committee on Antarctic Research によって承認されたデータ パブリッシャーとして GBIF に登録されているSCAR - Microbial Antarctic Resource System が、このリソースをパブリッシュしました。

キーワード

Metadata

連絡先

地理的範囲

Miers Valley, Antarctica

生物分類学的範囲

16S ssu rRNA

時間的範囲

収集方法

Aerosol samples were collected by filtration onto 0.2-μm-pore-size polycarbonate filters by impaction using solar-powered pumps (SKC, 224-PCXR8, Eighty Four, PA, USA) mounted 1 m above the ground in each location. Filters inserted into cassette apparatus but not exposed to air flow were used as controls. All filters and cassettes were UV-sterilized and rinsed with 70 % alcohol before use. Sampling apparatus was deployed 1 m above the ground surrounded with a 2 mm gauze baffle on the Miers Valley Floor (78°05′.78S, 163°47′.25E, approx. 270 m) and Miers Valley Ridge (78°06′.83S, 163°47′.18E, approx. 550 m).

Method step description:

1. Air sampling (see sampling description)
2. Filters were stored at −20 °C during transit from Antarctica and until processed.
3. Total DNA was extracted directly from the filters using the DNeasy Plant Mini Kit (Qiagen, CA, USA), after first washing with kit lysis buffer for 10 min. The remaining steps of the extraction were carried out according to the manufacturers instructions. Recovered DNA was quantified using NanoDrop™ (Thermo Scientific, Waltham, MA, USA).
4. For each sample, PCR targeting the V5–V7 region of the 16S rRNA gene was completed in duplicate. Each 30 μl reaction contained 1× PrimeSTAR buffer, 0.2 mM dNTPs, 0.75 U PrimeSTAR HS DNA Polymerase (Takara Holdings, Kyoto, Japan), 0.4 μM of primers Tx9 (5′-GGATTAGAWACCCBGGTAGTC-3′) and 1391R (5′-GACGGGCRGTGWGTRCA-3′), and 5 μl of template DNA. Thermal cycling conditions consisted of 94 °C for 3 min; 30 cycles of 94 °C for 20 s, 52 °C for 20 s, and 72 °C for 45 s; and 72 °C for 3 min. All thermal cycling was completed on a Bio-Rad DNA Engine Peltier Thermal Cycler 200 (Bio-Rad, Hercules, CA, USA). Duplicate reactions were pooled, and amplicons were size-selected from agarose gels using a MO BIO Gel Extraction Kit (MO BIO Laboratories, Carlsbad, CA, USA). Gel extracted products were cleaned using an Agencourt AMPure XP system (Beckman Coulter, Brea, CA, USA) and quantified using a Qubit dsDNA HS Assay Kit and Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA).
5. To prepare the amplicons for sequencing, a second round of PCR was completed in triplicate. PCR reactions were prepared as outlined above, but using 10 ng of purified amplicon as the template and primers MIDX-Tx9F (5′-CCATCTCATCCCTGCGTGTCTCCGACTCAG-MID-GGATTAGAWACCCBGGTAGTC-3′) and BacB_1391R (5′-CCTATCCCCTGTGTGCCTTGGCAGTCTCAG-GACGGGCRGTGWGTRCA-3′), adapted for one-way reads with unique MID identifiers for each sample. Thermal cycling conditions were as outlined above, but reduced to 13 cycles. Triplicate products were pooled, gel-extracted, cleaned, and quantified as outlined above, before quantification of amplifiable molecules using a KAPA Library Quantification Kit for Roche 454 Titanium/Universal (Kapa Biosystems, Woburn, MA, USA) on a Corbett Rotor-Gene 6000 real-time thermal cycler (Life Technologies). Amplicons were sequenced using the GS Junior Titanium emPCR Kit (Lib-L), the GS Junior Titanium Sequencing Kit, PicoTitrePlate Kit, and GS Junior System (Roche 454 Life Sciences, Branford, CT, USA) at The University of Waikato DNA Sequencing Facility.

書誌情報の引用

1. Bottos, E. M., Woo, A. C., Zawar-Reza, P., Pointing, S. B., & Cary, S. C. (2014). Airborne bacterial populations above desert soils of the McMurdo Dry Valleys, Antarctica. Microbial ecology, 67(1), 120-128.

追加のメタデータ