Figure 6. Generation of KO cell lines utilizing the sgRNA^KOLIBRY library.

(a) Schematic diagram of knockout cell line generation. (b) Schematic view of the genetic locus of the human TBKBP1 gene. The red arrow indicates the location of the target site of the sgRNA. The red sequence highlights the target site whereby the orange sequence indicates the PAM. (c) OutKnocker was used to analyze deep sequencing data obtained from gene targeting experiments. Each pie chart represents the deep sequencing data from an individual cell clone. Each pie piece sums up all reads that have the same indel mutation, with its size representing the proportion of total reads analyzed. Grey pie charts show reads without indel mutations, blue colors indicate indel mutations that are in frame and red colors indicate indel mutation reads that are out of frame, whereas the different shades of blue and red represent individual indel mutations in one pie chart. Depicted is an exemplary output for the generation of TBKBP1 knockout HEK cells. (d) Shown is a histogram summarizing the results of 19 individual gene targeting projects. Grey bars show the number of clones that carry only unmodified alleles, while purple bars indicate clones with at least one mutated allele and red bars clones with all-allelic frame shift mutations.