Table Of Content

LA Amaral-Zettler et al. – Supplemental information WebPanel 1. Sample collection and plastic identification We collected surface seawater using a clean flow-through seawater system, and water from 500- m depth using Niskin-bottles collected daily at noon during open-ocean transects in the Atlantic and Pacific (Figure 1b). We conducted our Atlantic transect across a latitudinal gradient over a 1- month timeframe, from mid-May (16 May 2012) through early June (11 Jun 2012), with the cooler temperate waters off the coast of Cape Cod, MA, being sampled in early June. For each water sample, we also simultaneously collected surface microplastics via neuston net tows with a 333-µm mesh size. Our study also included samples from sterile pieces of plastic immersed in seawater on the ship in the open Atlantic, and off docks in Woods Hole, MA; San Diego, CA; and St George’s, Grenada, in the Caribbean (WebTable 1). Sample sizes were as follows: (1) individual pieces of microplastic collected on shipboard via neuston net (n = 99); filter- concentrated seawater (n = 160); and substrates immersed in seawater on the ship or dockside (n = 87). Samples were preserved in lysis buffer and frozen at –20°C until DNA extraction. Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy was used to confirm and identify pieces that we were unable to characterize reliably using the Raman unit. A Perkin Elmer Spectrum 100 with a diamond-cell ATR attachment was used to make integrated surface measurements over a spot size diameter of ~1 mm. Polymer identification was determined by search and mixture techniques to match raw spectra with a library containing 500 polymers (Stadtler Spectral Database). Metadata associated with the samples, including identifications of the different plastics, are found in WebTable 1. DNA extraction, sequencing, and analyses Total genomic DNA was extracted from all samples using a commercial kit (Gentra Puregene Kit, Qiagen, Valencia, CA) following manufacturer’s instructions with the following modifications. Ready-Lyse™ lysozyme (10 µl of 1000 units µl–1 stock; Epicentre, Madison, WI) was added to each sample, followed by incubation at 37°C for 30 minutes. Proteinase K and 0.325 g of sterile 0.1-mm glass beads (MoBio, Carlsbad, CA) were added to each extraction tube, and tubes were vortexed at maximum speed for 1 minute using the MoBio Vortex Adapter (MoBio, Carlsbad, CA) to facilitate cell lysis. Extracts were stored at 4°C during experimental procedures and moved to –20°C for long-term storage. DNA amplicon next-generation sequencing determined bacterial community profiles of 346 samples. The V6 hypervariable region of 16S rRNA gene was amplified in triplicate with V6-specific primers, using a high-fidelity Taq polymerase (Platinum® Taq DNA Polymerase High Fidelity, Life Technologies, Grand Island, NY) as described in Eren et al. (2013). Pooled, triplicate amplification products were cleaned using MinElute columns (Qiagen, Valencia, CA) to remove excess primers and dimers. A second round of polymerase chain reaction (PCR) was performed, using 1–5 µl of first-round product as template, to add a barcode/index combination and Illumina adaptors to each amplicon, in preparation for paired-end sequencing on an Illumina HiSeq platform. The second-round reaction was run for ten cycles, using a high-fidelity Taq polymerase with the following cycling conditions: 94°C for 3 min; ten cycles of 94°C for 30 sec; 58°C for 45 sec; 72°C for 1 min; and a final extention at 72°C for 2 min. A negative template control was also run for each barcode/index combination. The final product size – including adaptors, index, barcode, and primers – was about 250 base pairs. Amplicons with visible product bands and clean negative controls were purified using AMPure XP beads (Agencourt, Beckman Coulter, Brea, CA) at a vol/vol ratio of 1.0, to remove primers and dimers. Following Ampure clean-up, the amplicons were quantified via a Quant-IT® PicoGreen™ assay (Invitrogen, Grand Island, NY) and pooled in equimolar concentration. Up to 192 pooled amplicons were combined with a metagenome at a 30:70 ratio (amplicons:metagenome) to be clustered and run in a single lane on the Illumina HiSeq platform. A total of 346 samples were sequenced on five different Illumina HiSeq runs. After sequencing, reads were de-multiplexed, and adaptor, barcode, and index sequences were removed. Paired-end reads were merged, retaining only pairs with complete overlap and no mismatches. For our samples, a total of 209 181 709 reads passed this stringent quality-filtering step. Across all 346 samples, the number of reads per sample retained after quality filtering averaged 604 571 reads per sample and a median of 447 726 reads per sample. Before analysis of the data, the number of reads per sample was normalized to the average value across all samples by random re-sampling. For those samples with less than the average number of reads, all reads were retained. We used the “usearch” algorithm implemented through QIIME (Caporaso et al. 2010) to cluster the normalized dataset into 89 865 clusters using the following criteria: a similarity of 97%, retaining clusters with a minimum size of 4, and no chimera-checking. Representative sequences from each cluster were selected using the “pick_rep_set.py” command in QIIME. Representative sequences were classified taxonomically using the GAST algorithm (Huse et al. 2008) and an in-house database of archaeal, bacterial, and eukaryotic V6 sequences (refv6.tgz, http://vamps.mbl.edu/resources/databases.php). QIIME outputs a matrix giving the number of occurrences of each cluster in each sample (ie the number of reads in each sample assigned to each cluster). The community matrix was first standardized by percent to correct for variation in sequencing depth, using the “decostand” function, with the option “total”. Bray-Curtis distances were calculated from the standardized matrix, using the function “vegdist” (Oksanen et al. 2012). NMDS analysis was run with the “metaMDS” function, using the Bray-Curtis distance matrix as input. Square root transformation of the data yielded comparable results so only non-transformed data were presented. Dispersion ellipses were drawn using the “ordiellipse” function. The dispersion ellipse represents standard deviation within each category of samples; the axis of the ellipse is determined by the correlation among samples. We examined our data for significant groupings by assigning samples to a “group” based on sample type (water, colonizations, or plastics), origin (Atlantic or Pacific), or polymer type (polyethylene, polypropylene, polystyrene, or polyethylene terephthalate) and tested the null hypothesis that there were no community-level differences between groups using a one-way Analysis of Similarity (ANOSIM) permutation test in PRIMER v6.1.16 (Clarke and Warwick 2001). Our ANOSIM tests used 999 random permutations of the data to assess the likelihood that the observed dissimilarity across a priori assigned groups occurred by chance. Observed species richness was obtained by randomly resampling the community matrix to 100 000 reads for each sample and counting the resulting operational taxonomic units (OTUs). We estimated species richness using the Chao and Jost constant-coverage estimator (Chao and Jost 2012). To calculate Chao-Jost Richness, we estimated the coverage for each sample using the software package iNEXT (Chao and Jost 2012; Hsieh et al. 2013), implemented in the R software package. The package iNEXT estimates coverage based on observed species richness (S ), the number of OTUs represented by one individual (f1), and the number of OTUs obs represented by two individuals (f2) (Chao and Jost 2012). The software also provides an interpolation of species richness versus coverage for “k” sample sizes, ranging from 1 to the actual sample size, and an extrapolation of species richness versus coverage for “k” sample sizes, ranging from (actual sample size + 1) to twice the actual sample size. The iNEXT function was run for each of our sampling sites, and the sites were sorted by estimated coverage. The goal was to obtain an estimate of species richness for each sampling site at a constant coverage value. To choose this coverage value, Chao and Jost (2012) recommended: (1) locating the sample with the lowest estimated coverage, (2) doubling the sample size, and (3) selecting the estimated coverage at twice the actual sample size from the extrapolated values for this sample. Estimated species richness for each sample was then calculated at this coverage value, using the output from iNEXT. For each sampling site, the interpolation and extrapolation values for species richness versus coverage were combined into a single data frame in R. The “lm” function in R was then used to estimate a slope and intercept for log(species richness) ~ coverage. Using the slope and intercept returned from the “lm” function, the expected richness at our selected coverage was calculated for each sample in the dataset. WebReferences Caporaso JG, Kuczynski J, Stombaugh J, et al. 2010. QIIME allows analysis of high-throughput community sequencing data. Nature Methods 7: 335–36. Chao A and Jost L. 2012. Coverage-based rarefaction and extrapolation: standardizing samples by completeness rather than size. Ecology 93: 2533–47. Clarke KR and Warwick RM. 2001. Change in marine communities: an approach to statistical analysis and interpretation (2nd edn). Plymouth, UK: PRIMER-E. Eren A, Vineis J, Morrison H, and Sogin M. 2013. A filtering method to generate high quality short reads using Illumina paired-end technology. PLoS ONE 8: e66643. Hsieh T, Ma K, and Chao A. 2013. iNEXT online: interpolation and extrapolation. http://chao.stat.nthu.edu.tw/blog/software-download. Viewed 5 Oct 2015. Huse SM, Dethlefsen L, Huber JA, et al. 2008. Exploring microbial diversity and taxonomy using SSU rRNA hypervariable tag sequencing. PLoS Genetics 4: e1000255. Oksanen J, Blanchet G, Kindt R, et al. 2012. Vegan: Community Ecology Package. R package version 1.17-3. WebPanel 2. Author contributions LAA-Z, ERZ, and TJM conceived the initial idea and contributed to experimental design and implementation. BS, GDB, and DWM performed experimental and molecular work. LAA-Z and BS performed bioinformatics analyses. CEM conducted Grenada sampling and acknowledges the assistance of J Brunet in doing so. LAA-Z, BS, ERZ, and TJM wrote the paper and all authors reviewed and edited the drafts. WebTable 1. MIMARKS Table with contextual and metadata Structured Comment Item Units COB_0066_20130716_Bv6 GRD_0020_2013_05_30_Bv6 GRD_0022_2013_05_30_Bv6 GRD_0023_2013_05_30_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation bacteria bacteria bacteria bacteria type experimental_factor experimental colonization colonization colonization colonization factor geo_loc_name geographic Pacific Grenada Grenada Grenada location (country and/or sea,region) lat_lon geographic +32.62083-‐117.12944 +11.99833-‐61.77167 +11.99833-‐61.77167 +11.99833-‐61.77167 location (latitude and longitude) collection_date collection date 2013-‐07-‐16 2013-‐05-‐30 2013-‐05-‐30 2013-‐05-‐30 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product manufactured product manufactured product manufactured product (feature) (ENVO:00003074) (ENVO:00003074) (ENVO:00003074) (ENVO:00003074) material environment polystyrene polystyrene glass glass (material) env_package environmental miscellaneous natural or miscellaneous natural or miscellaneous natural or miscellaneous natural or package artificial environment artificial environment artificial environment artificial environment samp_collect_device sample NA NA NA NA collection device or method samp_size amount or size L NA NA NA NA of sample collected nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit extraction Kit lib_reads_seqd library reads 1099910 192,339 7,165 32,385 sequenced lib_const_meth library Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification construction method target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target v6 v6 v6 v6 subfragment seq_meth sequencing Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq method pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C 20x(95°C for 30 sec, 60°C for 45 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for for 45 sec and 72°C for 45 sec and 72°C for 45 sec), 72°C 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), sec), 72°C for 5 min. 2nd for 5 min. 2nd step: 95°C for 3 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C step: 95°C for 3 min, min, 10x(95°C for 30 sec, 60°C for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 10x(95°C for 30 sec, 60°C for 45 sec and 72°C for 45 sec) 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 for 45 sec and 72°C for 45 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 quality check sop relevant MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC standard operating procedures url relevant http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ electronic resources depth depth m 0.5 0.5 0.5 0.5 temp temperature C na 28.1 28.1 28.1 salinity salinity ppt na na na na nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units GRD_0024_2013_05_30_Bv6 GRD_0027_2013_06_06_Bv6 GRD_0029_2013_06_06_Bv6 GRD_0031_2013_06_06_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor colonization colonization colonization colonization geo_loc_name geographic location (country and/or sea,region) Grenada Grenada Grenada Grenada lat_lon geographic location (latitude and longitude) +11.99833-‐61.77167 +11.99833-‐61.77167 +11.99833-‐61.77167 +11.99833-‐61.77167 collection_date collection date 2013-‐05-‐30 2013-‐06-‐06 2013-‐06-‐06 2013-‐06-‐06 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product manufactured product manufactured product manufactured product (feature) (ENVO:00003074) (ENVO:00003074) (ENVO:00003074) (ENVO:00003074) material environment (material) glass polypropylene high density polyethylene polystyrene env_package environmental miscellaneous natural or miscellaneous natural or miscellaneous natural or miscellaneous natural or package artificial environment artificial environment artificial environment artificial environment samp_collect_device sample collection device or method NA NA NA NA samp_size amount or size L of sample collected NA NA NA NA nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Kit Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit lib_reads_seqd library reads sequenced 95,282 448,979 113,620 95,508 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 45 sec and 72°C for 45 sec), 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 72°C for 5 min. 2nd step: 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 95°C for 3 min, 10x(95°C for 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 30 sec, 60°C for 45 sec and for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 72°C for 45 sec) 72°C for 5 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 min. 45 sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 0.5 0.5 0.5 0.5 temp temperature C 28.1 28.2 28.2 28.2 salinity salinity ppt na na na na nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units SEA_0013_20120516_Bv6 SEA_0014_20120516_Bv6 SEA_0019_20120517_Bv6 SEA_0020_20120517_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor water water water water geo_loc_name geographic location (country and/or sea,region) Atlantic Atlantic Atlantic Atlantic lat_lon geographic location (latitude and longitude) +18.67167-‐64.585 +18.67167-‐64.585 +20.03167-‐64.96667 +20.03167-‐64.96667 collection_date collection date 2012-‐05-‐16 2012-‐05-‐16 2012-‐05-‐17 2012-‐05-‐17 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment (feature) water water water water material environment saline water saline water saline water (material) saline water (ENVO:00002010) (ENVO:00002010) (ENVO:00002010) (ENVO:00002010) env_package environmental package water water water water samp_collect_device sample collection device or method 0.2 micron sterivex 0.2 micron sterivex 0.2 micron sterivex 0.2 micron sterivex samp_size amount or size L of sample collected 4 4 4 4 nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Gentra Puregene Yeast/Bac Kit Kit Kit Kit lib_reads_seqd library reads sequenced 777,593 446,474 1,131,785 838,417 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 72°C for 5 min. 2nd step: 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 95°C for 3 min, 10x(95°C for for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 30 sec, 60°C for 45 sec and 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 72°C for 45 sec) 72°C for 5 sec) 72°C for 5 min. 45 sec) 72°C for 5 min. 45 sec) 72°C for 5 min. min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 0 0 0 0 temp temperature C 27.6 27.6 26.9 26.9 salinity salinity ppt 36.26 36.26 36.21 36.21 nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units SEA_0022_20120517_Bv6 SEA_0026_20120518_Bv6 SEA_0029_20120518_Bv6 SEA_0031_20120519_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor waterDeep water microplastic water geo_loc_name geographic location (country and/or sea,region) Atlantic Atlantic Atlantic Atlantic lat_lon geographic location (latitude and longitude) +20.03167-‐64.96667 +21.52167-‐64.89167 +21.52167-‐64.89167 +22.30333-‐65.18 collection_date collection date 2012-‐05-‐17 2012-‐05-‐18 2012-‐05-‐18 2012-‐05-‐19 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product (feature) water water (ENVO:00003074) water material environment saline water saline water (material) (ENVO:00002010) (ENVO:00002010) high density polyethylene saline water (ENVO:00002010) env_package environmental miscellaneous natural or package water water artificial environment water samp_collect_device sample collection device or method 0.2 micron sterivex 0.2 micron sterivex 333 µm neuston net 0.2 micron sterivex samp_size amount or size L of sample collected 4 4 NA 4 nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Kit Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit lib_reads_seqd library reads sequenced 586,184 791,027 719,512 1,320,163 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 45 sec and 72°C for 45 sec), 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 72°C for 5 min. 2nd step: 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 95°C for 3 min, 10x(95°C for 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 30 sec, 60°C for 45 sec and for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 72°C for 45 sec) 72°C for 5 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 min. 45 sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 550 0 0 0 temp temperature C 12.3 25.8 25.8 25.9 salinity salinity ppt 35.59 36.40 36.40 36.00 nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units SEA_0032_20120519_Bv6 SEA_0035_20120519_Bv6 SEA_0036_20120519_Bv6 SEA_0037_20120520_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor water microplastic microplastic water geo_loc_name geographic location (country and/or sea,region) Atlantic Atlantic Atlantic Atlantic lat_lon geographic location (latitude and longitude) +22.30333-‐65.18 +22.30333-‐65.18 +22.30333-‐65.18 +23.28333-‐65.05167 collection_date collection date 2012-‐05-‐19 2012-‐05-‐19 2012-‐05-‐19 2012-‐05-‐20 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product manufactured product (feature) water (ENVO:00003074) (ENVO:00003074) water material environment saline water (material) (ENVO:00002010) high density polyethylene high density polyethylene saline water (ENVO:00002010) env_package environmental miscellaneous natural or miscellaneous natural or package water artificial environment artificial environment water samp_collect_device sample collection device or method 0.2 micron sterivex 333 µm neuston net 333 µm neuston net 0.2 micron sterivex samp_size amount or size L of sample collected 4 NA NA 4 nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Kit Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit lib_reads_seqd library reads sequenced 1,470,761 811,142 439,405 918,730 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 45 sec and 72°C for 45 sec), 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 72°C for 5 min. 2nd step: 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 95°C for 3 min, 10x(95°C for 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 30 sec, 60°C for 45 sec and for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 72°C for 45 sec) 72°C for 5 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 min. 45 sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 0 0 0 0 temp temperature C 25.9 25.9 25.9 26.0 salinity salinity ppt 36.00 36.00 36.00 36.01 nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units SEA_0038_20120520_Bv6 SEA_0041_20120520_Bv6 SEA_0042_20120520_Bv6 SEA_0043_20120521_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor water microplastic microplastic water geo_loc_name geographic location (country and/or sea,region) Atlantic Atlantic Atlantic Atlantic lat_lon geographic location (latitude and longitude) +23.28333-‐65.05167 +23.29167-‐65.08333 +23.29167-‐65.08333 +24.81167-‐64.57 collection_date collection date 2012-‐05-‐20 2012-‐05-‐20 2012-‐05-‐20 2012-‐05-‐21 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product manufactured product (feature) water (ENVO:00003074) (ENVO:00003074) water material environment saline water (material) (ENVO:00002010) polypropylene-‐isotactic polyethylene saline water (ENVO:00002010) env_package environmental miscellaneous natural or miscellaneous natural or package water artificial environment artificial environment water samp_collect_device sample collection device or method 0.2 micron sterivex 333 µm neuston net 333 µm neuston net 0.2 micron sterivex samp_size amount or size L of sample collected 4 NA NA 4 nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Kit Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit lib_reads_seqd library reads sequenced 1,013,061 735,804 674,663 503,956 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 45 sec and 72°C for 45 sec), 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 72°C for 5 min. 2nd step: 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 95°C for 3 min, 10x(95°C for 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 30 sec, 60°C for 45 sec and for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 72°C for 45 sec) 72°C for 5 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 min. 45 sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 0 0 0 0 temp temperature C 26.0 26.0 26.0 26.2 salinity salinity ppt 36.01 36.01 36.01 36.07 nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM Structured Comment Item Units SEA_0044_20120521_Bv6 SEA_0049_20120521_Bv6 SEA_0051_2012_05_22_Bv6 SEA_0052_2012_05_22_Bv6 Name project_name project name Plastisphere Plastisphere Plastisphere Plastisphere investigation_type investigation type bacteria bacteria bacteria bacteria experimental_factor experimental factor water microplastic water water geo_loc_name geographic location (country and/or sea,region) Atlantic Atlantic Atlantic Atlantic lat_lon geographic location (latitude and longitude) +24.81167-‐64.57 +24.82833-‐64.58167 +26.07-‐64.18667 +26.07-‐64.18667 collection_date collection date 2012-‐05-‐21 2012-‐05-‐21 2012-‐05-‐22 2012-‐05-‐22 biome environment marine biome marine biome marine biome marine biome (biome) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) (ENVO:00000428) feature environment manufactured product (feature) water (ENVO:00003074) water water material environment saline water (material) (ENVO:00002010) high density polyethylene saline water (ENVO:00002010) saline water (ENVO:00002010) env_package environmental miscellaneous natural or package water artificial environment water water samp_collect_device sample collection device or method 0.2 micron sterivex 333 µm neuston net 0.2 micron sterivex 0.2 micron sterivex samp_size amount or size L of sample collected 4 NA 4 4 nucl_acid_ext nucleic acid Gentra Puregene Yeast/Bac Gentra Puregene Yeast/Bac extraction Kit Kit Gentra Puregene Yeast/Bac Kit Gentra Puregene Yeast/Bac Kit lib_reads_seqd library reads sequenced 1,643,710 481,792 48,195 73,925 lib_const_meth library construction method Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification Fusion 2-‐step amplification target_gene target gene 16S rRNA 16S rRNA 16S rRNA 16S rRNA target_subfragment target subfragment v6 v6 v6 v6 seq_meth sequencing method Illumina HiSeq Illumina HiSeq Illumina HiSeq Illumina HiSeq pcr_primers pcr primers Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) Forward Primers (967F) CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CTAACCGANGAACCTYACC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CNACGCGAAGAACCTTANC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC CAACGCGMARAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC ATACGCGARGAACCTTACC Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) Reverse Primer (1064R) CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT CGACRRCCATGCANCACCT pcr_cond pcr conditions 1st step: 95°C for 3 min, 20x(95°C for 30 sec, 60°C for 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 1st step: 95°C for 3 min, 45 sec and 72°C for 45 sec), 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 20x(95°C for 30 sec, 60°C for 72°C for 5 min. 2nd step: 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 45 sec and 72°C for 45 sec), 95°C for 3 min, 10x(95°C for 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 72°C for 5 min. 2nd step: 95°C 30 sec, 60°C for 45 sec and for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, for 3 min, 10x(95°C for 30 sec, 72°C for 45 sec) 72°C for 5 60°C for 45 sec and 72°C for 60°C for 45 sec and 72°C for 45 60°C for 45 sec and 72°C for 45 min. 45 sec) 72°C for 5 min. sec) 72°C for 5 min. sec) 72°C for 5 min. seq_quality_check sequence quality check PMID: 23799126 PMID: 23799126 PMID: 23799126 PMID: 23799126 sop relevant standard operating procedures MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC MBL-‐JBPC url relevant electronic resources http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ http://vamps.mbl.edu/ depth depth m 0 0 0 0 temp temperature C 26.2 26.2 26.0 26.0 salinity salinity ppt 36.07 36.07 36.07 36.07 nitrate nitrate µM nitrite nitrite µM ammonium ammonium µM phosphate phosphate µM silicate silicate µM

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2008) and an in-house database of archaeal, bacterial, and eukaryotic V6 sequences (refv6.tgz, http://vamps.mbl.edu/resources/databases.php).

Amaral-Zettler WOM PDF

90 Pages·2015·2.33 MB·English

by Linda Amaral Zettler

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by Linda Amaral Zettler| 2015| 90 pages| 2.33| English

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2008) and an in-house database of archaeal, bacterial, and eukaryotic V6 sequences (refv6.tgz, http://vamps.mbl.edu/resources/databases.php).

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Author:	Linda Amaral Zettler
Publication Year:	2015
Pages:	90
Language:	English
File Size:	2.33
Format:	PDF
Price:	FREE

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