16S rRNA sequencing of human lung metagenome: sample Pa02_003 - SRA

SRX472699: 16S rRNA sequencing of human lung metagenome: sample Pa02_003
1 LS454 (454 GS FLX Titanium) run: 16,505 spots, 9M bases, 17.9Mb downloads

Design: Construction protocol: The v6-v4 hypervariable regions of the SSU rRNA gene were amplified using degenerate primers. The primers were 1064R (CGACRRCCATGCANCACCT) and 518F (CCAGCAGCYGCGGTAAN) fused to Roche GSFLX amplicon sequencing adapters and including 5 nt multiplexing barcodes. We generated PCR amplicons in triplicate 33 uL reaction volumes with an amplification cocktail containing 1.0 U Platinum Taq Hi-Fidelity Polymerase (Invitrogen, Carlsbad, CA), 1X Hi-Fidelity buffer, 200 uM dNTP PurePeak DNA polymerase mix (Pierce Nucleic Acid Technologies, Milwaukee, WI), 1.5 mM MgSO4 and 0.2 uM of each primer. We added approximately 10-25 ng template DNA to each PCR and ran a no-template control for each primer pair. Amplification conditions were: initial 94C, 3 minute denaturation step; 30 cycles of 94C for 30s, 60C for 60s, and 72C for 90s; final 10 minute extension at 72C. The triplicate PCR reactions were pooled after amplification and purified using Ampure beads. Purified DNA was eluted in 30 uL of Qiagen buffer EB. We assessed the quality, size and concentration of PCR products on a Perkin Elmer Caliper GX. Reads were demultiplexed and barcodes removed for submission.

Submitted by: Geisel School of Medicine at Dartmouth

Study: Microbial community analysis of sputum samples from adults and pediatric patients

PRJNA237382 • SRP038106 • All experiments • All runs

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The goal of the study is to understand the similarities and differences in the microbial communities in adults and pediatric patients

Sample: Survey related sample for polymicrobial

SAMN02628442 • SRS559134 • All experiments • All runs

Organism: human lung metagenome

Library:

Name: Pa02_003

Instrument: 454 GS FLX Titanium

Strategy: AMPLICON

Source: METAGENOMIC

Selection: PCR

Layout: SINGLE

Spot descriptor:

5 forward

Runs: 1 run, 16,505 spots, 9M bases, 17.9Mb

Run	# of Spots	# of Bases	Size	Published
SRR1171620	16,505	9M	17.9Mb	2014-04-15

ID:: 653189

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