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Sample GSM2330268 Query DataSets for GSM2330268
Status Public on Sep 18, 2020
Title environmental_sample_pool_A
Sample type SRA
 
Source name environmental samples that include metal working fluid and air from a machine facility
Organism uncultured bacterium
Characteristics linkerprimersequence: CCGGACTACHVGGGTWTCTAAT
Extracted molecule genomic DNA
Extraction protocol ion exchange column (Qiagen)
Amplicon library preparation was performed using an automated platform (Biomek 4000) using a custom liquid handling method. For each sample, the V4 region of the bacterial 16S rRNA gene was amplified in duplicate reactions, using primer set 515F/806R, which nearly universally amplifies bacterial and archaeal 16S rRNA genes.1,2 Each unique barcoded amplicon was generated in pairs of 25μl reactions with the following reaction conditions: 11μl PCR-grade H2O, 10μl Hot MasterMix (5 Prime Cat# 2200410), 2μl of forward and reversed barcoded primer (5μM) and 2μl template DNA. Reactions were run on a C1000 Touch Thermal Cycler (Bio-Rad) with the following cycling conditions: initial denaturing at 94°C for 3 min followed by 35 cycles of denaturation at 94°C for 45 seconds, annealing at 58°C for 1 minute, and extension at 72 C for 90 seconds, with a final extension of 10 min at 72°C. Amplicons were quantified using Agilent 2200 TapeStation system and pooled. Purification was then performed using Ampure XT (Beckman Coulter Cat# A63882) as per the manufacturer instructions
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina MiSeq
 
Description ribosomal DNA
The SRR for this sample (SRR4298944) contains data for the forward and reverse strand as well as the barcode information in fastq format. The three separate fastq files can be obtained with the command: fastq-dump --split-files SRR4298944.
Data processing Library strategy: DNA-seq
demultiplexing and filtering of short (<150 nt) and low quality reads. Barcode information is available in the file "Sample_barcode_metadata.txt" on the series record.
de novo clustering of the sequences into operational taxonomic units (OTUs) with the UCLUST program using a 97% similarity threshold
taxonomical assignment of each OTU by running the RDP Classifier(8) at 80% bootstrap confidence on a selected representative sequence from each OTU;
alignment of representative sequences using PyNAST(9) with the Greengenes core-set alignment template
phylogenetic tree reconstruction from the representative sequences for each OTUs using the FASSTTREE program
Bray Curtis distance calculations
Supplementary_files_format_and_content: read abundance for OTUs. The file Sample_barcode_metadata.txt.gz on the series record contains the barcode information and sample source for all raw data contained in the multiplexed raw data file.
 
Submission date Sep 27, 2016
Last update date Sep 18, 2020
Contact name Leopoldo Nicolas Segal
E-mail(s) Leopoldo.Segal@nyumc.org
Phone 2122636479
Organization name NYU School of Medicine
Department Medicine
Street address 462 First Avenue NBV 6N5-6N8
City New York
State/province NY
ZIP/Postal code 10016
Country USA
 
Platform ID GPL22493
Series (1)
GSE87406 Possible role of environmental microbiome on B-cell bronchiolitis-alveolar ductitis and emphysema in metal machining workers
Relations
BioSample SAMN05830450
SRA SRX2193539

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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