title: Amplicon Sequencing of gRNAs from Druggable Genome CRISPR mutagenized Huh7.5.1 cells after live/dead selection with DENV2
design: Amplicon Sequencing of gRNAs from Druggable Genome CRISPR mutagenized Huh7.5.1 cells after live/dead selection with DENV2
library_construction_protocol: >1000-fold coverage of mutagenized cells was infected with DENV2 at MOI 0.1. Resistant cell colonies were harvested. Uninfected starting populations of mutagenized cells were used as the unselected reference. Total genomic DNA from both virus-resistant and uninfected cells was respectively extracted using QIAamp DNA Mini Kit (Qiagen). A first round of PCR was used to amplify the guide RNA sequences encoded in the gDNA, followed by a second round of PCR to add the barcodes/adapters for amplicon sequencing. 2% agarose gels and a QIAquick gel extraction kit were used to purify the amplicons.