SRX2850175: Amplicon seq: human sino-nasal metagenome
1 PACBIO_SMRT (PacBio RS II) run: 1 spots, 544 bases, 51,525b downloads
Design: Full-length 16S amplifications were performed using: 1μl of total DNA as template; 0.25 μM of the universal 16S primers F27 and R1492 with four different sets of asymmetric barcodes; and GoTaq Hot Start Master Mix (Promega) in a 50μl final volume. Cycling conditions were: 94 °C, 3 min; 35 cycles of 94 °C 30 sec, 54 °C 30 sec, 72 °C 2 min; following by a 5 min final elongation at 72 °C. PCR products were cleaned with AxyPrepTM MagPCR (Corning Life Sciences) according to the manufacturer’s protocol and eluted in 40μl of water. Cleaned PCR products were quantified using the Bio-Rad QX200 droplet digital PCR (Bio-Rad) and QX200 EvaGreen® Supermix with primers F357 and R534 targeting the V3 variable region of 16S rDNA. Based on the results, amplicon libraries were normalized to the same concentration prior to pooling. Pooling was always performed using amplicon libraries with distinct barcodes. Multiplexing was performed with 2-4 libraries per pool.
Submitted by: Joshua Earl (Drexel1)
Study:
16s rRNA sequencing of sino-nasal microbiome and mock-community samplesshow Abstracthide AbstractAnalysis of 16s rRNA reads from PacBio sequencing using a custom made analysis pipeline. 2 mock communities (BEI and JGI) and a set of sino-nasal samples were sequenced as a part of this project.
Library:
Name: negN4_control
Instrument: PacBio RS II
Strategy: AMPLICON
Source: METAGENOMIC
Selection: PCR
Layout: SINGLE
Runs:
1 run, 1 spots, 544 bases, 51,525b