title: OutKnocker: a web tool for rapid and simple genotyping of designer nuclease edited cell lines.
abstract: The application of designer nucleases allows the induction of DNA double-strand breaks (DSBs) at user-defined genomic loci. Due to imperfect DNA repair mechanisms, DSBs can lead to alterations in the genomic architecture, such as the disruption of the reading frame of a critical exon. This can be exploited to generate somatic knockout cell lines. While high genome editing activities can be achieved in various cellular systems, obtaining cell clones that contain all-allelic frameshift mutations at the target locus of interest remains a laborious task. To this end, we have developed an easy-to-follow deep sequencing workflow and the evaluation tool OutKnocker (www.OutKnocker.org), which allows convenient, reliable, and cost-effective identification of knockout cell lines.
authors.affiliation: Institute of Molecular Medicine, University Hospital, University of Bonn, 53127 Bonn, Germany;
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Schmid-Burgk
authors.affiliation: Institute of Molecular Medicine, University Hospital, University of Bonn, 53127 Bonn, Germany;
authors.forename: Tobias
authors.initials: T
authors.lastname: Schmidt
authors.affiliation: Institute of Molecular Medicine, University Hospital, University of Bonn, 53127 Bonn, Germany;
authors.forename: Moritz M
authors.initials: MM
authors.lastname: Gaidt
authors.affiliation: Institute of Innate Immunity, University Hospital, University of Bonn, 53127 Bonn, Germany.
authors.forename: Karin
authors.initials: K
authors.lastname: Pelka
authors.affiliation: Institute of Innate Immunity, University Hospital, University of Bonn, 53127 Bonn, Germany.
authors.forename: Eicke
authors.initials: E
authors.lastname: Latz
authors.affiliation: Institute of Molecular Medicine, University Hospital, University of Bonn, 53127 Bonn, Germany;
authors.forename: Thomas S
authors.initials: TS
authors.lastname: Ebert
authors.affiliation: Institute of Molecular Medicine, University Hospital, University of Bonn, 53127 Bonn, Germany; veit.hornung@uni-bonn.de.
authors.forename: Veit
authors.initials: V
authors.lastname: Hornung
mesh_terms: D045744:Cell Line, Tumor; D004720:Endonucleases / Q000378:metabolism*; D016368:Frameshift Mutation; D055786:Gene Knockout Techniques; D060005:Genotyping Techniques / Q000191:economics / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D017393:RNA Editing; D017422:Sequence Analysis, DNA / Q000191:economics / Q000379:methods*; D012984:Software; D064878:Web Browser
chemical_list: D004720:Endonucleases