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URL: https://dx.doi.org/10.17504/protocols.io.nuvdew6

Authors: Linda A. Amaral-Zettler, Markus Bauer, Donna Berg-Lyons, Jason Betley, J. Greg Caporaso, Hugh W. Ducklow, Noah Fierer, Louise Fraser, Jack A. Gilbert, Niall Gormley, James Huntley, Susan M. Huse, Janet K. Jansson, Simon N. Jarman, Rob Knight, Chris L. Lauber, Elizabeth A. McCliment, Sarah M. Owens, Geoff Smith, Luke Thompson, Hege Vestheim, William A. Walters

Group: Earth Microbiome Project

Summary: The 18S protocol detailed here is designed to amplify eukaryotes broadly with a focus on microbial eukaryotic lineages. The primers target the 18S SSU rRNA and are based on those of Amaral-Zettler et al. (2009). The constructs are designed to be used with the Illumina platform.For running these libraries on the MiSeq and HiSeq, please make sure you read the supplementary methods of Caporaso et al. (2012). You will need to make your sample more complex by adding 5-10% PhiX to your run.The outlines of the protocol are the same as the 16S protocol, but different primers, PCR conditions, and sequencing primers are used. In addition, we have designed a blocking primer that reduces the amplification of vertebrate host DNA to be used on host-associated samples, especially those that have a low eukaryotic biomass. Blocking primer strategy is based on Vestheim et al. (2008).

Associated Publications: Ul-Hasan S, Bowers RM, Figueroa-Montiel A, Licea-Navarro AF, Beman JM, Woyke T, Nobile CJ (2019) Community ecology across bacteria, archaea and microbial eukaryotes in the sediment and seawater of coastal Puerto Nuevo, Baja California. PLoS ONE 14(2): e0212355. doi: 10.1371/journal.pone.0212355

Affiliations: EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium, EMP Consortium

External URL: http://press.igsb.anl.gov/earthmicrobiome/protocols-and-standards/18s/

Version: 1

Publication Date: 2018

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