DNA was extracted from ~100 mg of stool using enzymatic treatment with 15 mg/ml lysozyme and proteinase K (Qiagen) for 10 min incubation followed by mechanical lysis using 0.1mm glass beads and 3 min bead beating on the “homogenize” setting of a Mini Bead beater-8 (Biospec Products). After lysis, samples were centrifuged for 5 minutes at 30,000 rpm to pellet debris and supernatant passed through a QIAshredder spin column (Qiagen) prior to proceeding with nucleic acid extraction using a Allprep DNA/RNA Mini Kit (Qiagen) following manufacturer protocol. Metagenomic libraries were constructed from 100–250 pg of DNA using the Nextera XT DNA Library Preparation Kit (Illumina) as per the manufacturer’s recommended protocol. Sequencing was performed on the HiSeq 2500 2× 101 PE platform (Illumina), targeting ~2.5 Gb of sequence per aliquot. Demultiplexing, BAM and FASTQ file generation were performed using the Picard suite (https://broadinstitute.github.io/picard ).
Hiseq 2500 2 101 pe platform
Manufactured by Illumina
The HiSeq 2500 2× 101 PE platform is a high-throughput sequencing system designed for DNA and RNA analysis. It supports paired-end sequencing with read lengths of up to 101 base pairs. The system is capable of generating large amounts of sequence data in a single run.
Automatically generated - may contain errors
2 protocols using hiseq 2500 2 101 pe platform
1
Metagenomic DNA Extraction from Stool
2
Stool DNA Extraction and Metagenomic Sequencing
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DNA was extracted from ~100 mg of stool using enzymatic treatment with 15 mg/ml lysozyme and proteinase K (Qiagen) for 10 min incubation followed by mechanical lysis using 0.1mm glass beads and 3 min bead beating on the "homogenize" setting of a Mini Bead beater-8 (Biospec Products). After lysis, samples were centrifuged for 5 minutes at 30,000 rpm to pellet debris and supernatant passed through a QIAshredder spin column (Qiagen) prior to proceeding with nucleic acid extraction using a Allprep DNA/RNA Mini Kit (Qiagen) following manufacturer protocol. Metagenomic libraries were constructed from 100-250 pg of DNA using the Nextera XT DNA Library Preparation Kit (Illumina) as per the manufacturer's recommended protocol. Sequencing was performed on the HiSeq 2500 2× 101 PE platform (Illumina), targeting ~2.5 Gb of sequence per aliquot. Demultiplexing, BAM and FASTQ file generation were performed using the Picard suite (https://broadinstitute.github.io/picard).
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