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

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

Amaral-Zettler WOM PDF

90 Pages·2015·2.33 MB·English
by  Linda Amaral Zettler
Save to my drive
Quick download
Download
Upgrade Premium
Most books are stored in the elastic cloud where traffic is expensive. For this reason, we have a limit on daily download.

Preview Amaral-Zettler WOM

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

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

The list of books you might like

Upgrade Premium
Most books are stored in the elastic cloud where traffic is expensive. For this reason, we have a limit on daily download.
DMCA & Copyright: Dear all, most of the website is community built, users are uploading hundred of books everyday, which makes really hard for us to identify copyrighted material, please contact us if you want any material removed.
Copyright @ PDFdrive.to, 2022 | We love our users