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The objective of this study was to evaluate the most effective method of DNA extraction of oral mouthwash samples for use in microbiome studies that utilize next generation sequencing (NGS). Eight enzymatic and mechanical DNA extraction methods were tested. Extracted DNA was amplified using barcoded primers targeting the V6 variable region of the bacterial 16S rRNA gene and the ITS1 region of the fungal ribosomal gene cluster and sequenced using the Illumina NGS platform. Sequenced reads were analyzed using QIIME and R. The eight methods yielded significantly different quantities of DNA (p < 0.001), with the phenol-chloroform extraction method producing the highest total yield. There were no significant differences in observed bacterial or fungal Shannon diversity (p = 0.64, p = 0.93 respectively) by extraction method. Bray-Curtis beta-diversity did not demonstrate statistically significant differences between the eight extraction methods based on bacterial (R2 = 0.086, p = 1.00) and fungal (R2 = 0.039, p = 1.00) assays. No differences were seen between methods with or without bead-beating. These data indicate that choice of DNA extraction method affect total DNA recovery without significantly affecting the observed microbiome.
Pubmed ID: 30728424
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Software program to align Illumina reads, optionally with PCR primers embedded in the sequence, and reconstruct an overlapping sequence.
View all literature mentionsA publicly available tool that is able to filter, reformat and trim your genomic and metagenomic sequence data and provide you summary statistics for your sequence data. The interactive web interface facilitates visualizations of the results and export functionality for subsequent data processing. The standalone lite version is written in Perl and does not require any non-core Perl modules. The lite version is primarily designed for data preprocessing and does not generate summary statistics in graphical form.
View all literature mentionsTHIS RESOURCE IS NO LONGER IN SERVICE. Documented on February 23,2023.Software package for comparison and analysis of microbial communities, primarily based on high-throughput amplicon sequencing data, but also supporting analysis of other types of data. QIMME analyzes and transforms raw sequencing data generated on Illumina or other platforms to publication quality graphics and statistics.
View all literature mentionsSource code that infers approximately-maximum-likelihood phylogenetic trees from alignments of nucleotide or protein sequences. It uses the Jukes-Cantor or generalized time-reversible (GTR) models of nucleotide evolution and the JTT, WAG, or LG models of amino acid evolution.
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