Marker gene sequencing of sample LSM_S5c_Mid - SRA

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SRX23952978: Marker gene sequencing of sample LSM_S5c_Mid
1 ILLUMINA (Illumina MiSeq) run: 125,049 spots, 62.8M bases, 34.4Mb downloads

Design: The v4-v5 hypervariable regions of the bacterial SSU rRNA gene were amplified using degenerate primers. The primers were 518F (CCAGCAGCYGCGGTAAN) and 926R (CCGTCAATTCNTTTRAGT CCGTCAATTTCTTTGAGT CCGTCTATTCCTTTGANT). Amplification was done with fusion primers containing the 16S-only sequences fused to Illumina adapters. The forward primers included a 5 nt multiplexing barcode and the reverse a 6 nt index. We generated PCR amplicons in triplicate 33 uL reaction volumes with an amplification cocktail containing 0.67 U SuperFi Taq Polymerase (Invitrogen, Carlsbad, CA), 1X enzyme buffer (includes MgCl2), 200 uM dNTP PurePeak DNA polymerase mix (ThermoFisher), and 0.3 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, 57C for 45s, and 72C for 60s; final 2 minute extension at 72C. The triplicate PCR reactions were pooled after amplification, visualized with the negative controls on a Caliper LabChipGX or Agilent TapeStation 4200, and purified using Ampure followed by PicoGreen quantitation and Ampure size selection.

Submitted by: Marine Biological Laboratory

Study: Little Sippewissett Marsh sediment cores Targeted loci environmental

PRJNA1088085 • SRP495270 • All experiments • All runs

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Samples were collected in October, 2015 as part of a University of Chicago field course, Experimental Biology by the Sea: Marine Microbial Ecology. DNA extracted from the samples served as template for 16s marker gene amplification and sequencing to determine bacterial community composition. Course instructors, Julie A. Huber WHOI, Karl Matlin UChicago, Linda Amaral-Zettler NIOZ.

Sample:

SAMN40460097 • SRS20753474 • All experiments • All runs

Organism: salt marsh metagenome

Library:

Name: LSM_S5c_Mid_Bv4v5

Instrument: Illumina MiSeq

Strategy: AMPLICON

Source: METAGENOMIC

Selection: PCR

Layout: PAIRED

Runs: 1 run, 125,049 spots, 62.8M bases, 34.4Mb

Run	# of Spots	# of Bases	Size	Published
SRR28346445	125,049	62.8M	34.4Mb	2024-03-14

ID:: 32258267

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