16S amplification of homo sapiens: gut metagenome - SRA

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SRX1442359: 16S amplification of homo sapiens: gut metagenome
1 ILLUMINA (Illumina MiSeq) run: 12,045 spots, 4M bases, 2.1Mb downloads

Design: 16S rRNA gene was amplified from DNA extracted from patient fecal samples and sequenced on an llumina MiSeq platform. Replicate PCR reactions were performed using modified universal bacterial primers designed to amplify the V4-V5 16S rRNA region: 563F (59-nnnnnnnn-NNNNNNNNNNNN-AYTGGGYDTAAAGN G-39) and 926R (59- nnnnnnnn-NNNNNNNNNNNN-CCGTCAATTYHTTTR AGT-39). Each reaction contained 50 ng of purified DNA, 0.2 mM dNTPs, 1.5 uM MgCl2, 1.25 U Platinum TaqDNA polymerase, 2.5 ul of 10x PCR buffer, and 0.2 uM of each primer. A unique 12-base Golay barcode (Ns) preceded the primers for sample identification after pooling amplicons. One to eight additional nucleotides were added before the barcode to offset the sequencing of the primers. Cycling conditions were the following: 94 ĄC for 3 min, followed by 27 cycles of 94 ĄC for 50 s, 51 ĄC for 30 s, and 72 ĄC for 1 min, where the final elongation step was performed at 72 ĄC for 5 min. Replicate PCRs were combined and were subsequently purified using the Qiaquick PCR Purification Kit (Qiagen). Using the Illumina TruSeq Sample Preparation procedure, PCR products were quantified and pooled at equimolar amounts before Illumina barcodes and adaptors were ligated on. The completed library was sequenced on an Ilumina Miseq platform according to the Illumina recommended protocol.

Submitted by: MSK

Study: Human gut Metagenome

PRJNA302832 • SRP066514 • All experiments • All runs

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16S ribosomal RNA (rRNA) gene amplicon sequencing of fecal samples collected from melanoma patients prior to ipilimumab treatment

Sample: !6S_Illumina_MiSeq_fecal_sample_pt_PtC_31

SAMN04281049 • SRS1172101 • All experiments • All runs

Organism: human gut metagenome

Library:

Name: Sample31

Instrument: Illumina MiSeq

Strategy: AMPLICON

Source: METAGENOMIC

Selection: PCR

Layout: PAIRED

Runs: 1 run, 12,045 spots, 4M bases, 2.1Mb

Run	# of Spots	# of Bases	Size	Published
SRR2940935	12,045	4M	2.1Mb	2015-11-26

ID:: 2040029

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