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**Aim 3: Establish culture collection of microbial eukaryotes associated with Zostera marina in Bodega Bay, California.**  _Objectives:_ Environmental molecular barcoding studies offer the benefit of high-throughput biodiversity discovery, especially of organisms that are recalcitrant to laboratory culture. However, they suffer from biases during DNA extraction and PCR, and can exclude important, even dominant, taxa**(REF)** maybe  \cite{HoefEmden_2012}or this 16S paper  \cite{Baker_2003}. They also fall short of providing the means to link biodiversity discovery with functional, ecological significance. The value of DNA sequencing-based studies can be improved by augmenting them with a rich, phylogenetically diverse culture collection. Such collections enable the sequencing of isolate genomes, providing information about metabolic potential, as well as reference sequences for shotgun metagenomics. They also offer a resource for future experimental work. Culturing diverse microbial eukaryotes from seagrass beds is a labor-intensive process, requiring much time and multiple sampling efforts returning fresh samples. Therefore, we will focus our initial efforts on Zostera marina beds in Bodega Bay, California, where we have full access to the Bodega Marine Laboratory and the required permits for collection. All isolates will be characterized using both microscopy and molecular methods (described below). _Culturing/Isolation_: We broadly define microbial eukaryotes to include both single-celled and multicellular eukaryotic organisms that are invisible to the naked eye, spanning every major eukaryotic lineage. These represent tremendous metabolic diversity, including photosynthetic and non-photosynthetic, obligate anaerobes and anaerobes, free-living and parasitic. Therefore, we will employ a wide variety of culture techniques to isolate them, including single cell sorting and enrichment cultures using seagrass rhizosphere sediments, filtered bulk seawater, and seagrass tissues as as sources (see \cite{Torta_2014}). We will use an isolation/characterization pipeline published by our lab \cite{Dunitz_2015} and focus on variety of media recipes and culture techniques to maximize diversity discovery \cite{Lee_1992}. We will also target specific taxa known to play important roles in seagrass health \cite{Muehlstein_1991} and important ecosystem services such as decomposition \cite{Newell_1996}, the carbon cycle \cite{Smetacek_1999} and primary production \cite{Field_1998} \cite{Armbrust_2009} \cite{Higgins_2012}. We have had successful experience in our lab culturing diverse amoeba, fungi, and ciliates.