Titlebook: Next Generation Sequencing; Methods and Protocol Steven R. Head,Phillip Ordoukhanian,Daniel R. Salo Book 2018 Springer Science+Business Med

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Hi-Plex for Simple, Accurate, and Cost-Effective Amplicon-based Targeted DNA Sequencing,techniques 58:33–36, 2015). At its core is the principle of using gene-specific primers (GSPs) to “seed” (or target) the reaction and universal primers to “drive” the majority of the reaction. In this manner, effects on amplification efficiencies across the target amplicons can, to a large extent, b

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ClickSeq: Replacing Fragmentation and Enzymatic Ligation with Click-Chemistry to Prevent Sequence Cick-chemistry in place of enzymes for the ligation of sequencing adaptors. We found that this approach dramatically reduces artifactual chimera formation, allowing the study of rare recombination events that include viral replication intermediates and defective-interfering viral RNAs. ClickSeq illus

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Genome-Wide Analysis of DNA Methylation in Single Cells Using a Post-bisulfite Adapter Tagging Apprlfite-based approaches, it is possible to obtain comprehensive genome-wide maps of the mammalian DNA methylation landscape with a single-nucleotide resolution and absolute quantification. However, these methods were only applicable to bulk populations of cells. Here, we present a protocol to perform

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Sampling, Extraction, and High-Throughput Sequencing Methods for Environmental Microbial and Viral tudy of as-yet uncultured microbes and viruses. Studies of complex microbial communities require high-quality data to generate valid results. Here, we detail current methods of microbial and viral community sample acquisition, DNA extraction, sample preparation, and sequencing on Illumina high-throu

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A Bloody Primer: Analysis of RNA-Seq from Tissue Admixtures,terogeneous tissues, cell admixtures with relative proportions that can vary several fold across samples, poses a significant challenge. Blood is perhaps the most egregious example. Here, we describe in detail a computational pipeline for RNA-Seq data preparation and statistical analysis, with devel

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Next-Generation Sequencing of Genome-Wide CRISPR Screens,a powerful tool for systematic genomic perturbation in mammalian cells and provide an alternative to previous screens utilizing RNA interference technology. The wide availability of these libraries through public plasmid repositories as well as the decreasing cost and speed in quantifying these scre

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