title: Genome-wide transposon screening and quantitative insertion site sequencing for cancer gene discovery in mice.
abstract: Transposon-mediated forward genetics screening in mice has emerged as a powerful tool for cancer gene discovery. It pinpoints cancer drivers that are difficult to find with other approaches, thus complementing the sequencing-based census of human cancer genes. We describe here a large series of mouse lines for insertional mutagenesis that are compatible with two transposon systems, PiggyBac and Sleeping Beauty, and give guidance on the use of different engineered transposon variants for constitutive or tissue-specific cancer gene discovery screening. We also describe a method for semiquantitative transposon insertion site sequencing (QiSeq). The QiSeq library preparation protocol exploits acoustic DNA fragmentation to reduce bias inherent to widely used restriction-digestion-based approaches for ligation-mediated insertion site amplification. Extensive multiplexing in combination with next-generation sequencing allows affordable ultra-deep transposon insertion site recovery in high-throughput formats within 1 week. Finally, we describe principles of data analysis and interpretation for obtaining insights into cancer gene function and genetic tumor evolution.
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Mathias J
authors.initials: MJ
authors.lastname: Friedrich
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: Rad
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Iraad F
authors.initials: IF
authors.lastname: Bronner
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Alexander
authors.initials: A
authors.lastname: Strong
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.|German Cancer Research Center (DKFZ), Heidelberg, &German Cancer Consortium (DKTK), Heidelberg, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Weber
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Matthew
authors.initials: M
authors.lastname: Mayho
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Hannes
authors.identifier: http://orcid.org/0000-0001-7573-1703
authors.initials: H
authors.lastname: Ponstingl
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.|German Cancer Research Center (DKFZ), Heidelberg, &German Cancer Consortium (DKTK), Heidelberg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Engleitner
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Carolyn
authors.initials: C
authors.lastname: Grove
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.|German Cancer Research Center (DKFZ), Heidelberg, &German Cancer Consortium (DKTK), Heidelberg, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Pfaus
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.|German Cancer Research Center (DKFZ), Heidelberg, &German Cancer Consortium (DKTK), Heidelberg, Germany.
authors.forename: Dieter
authors.initials: D
authors.lastname: Saur
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.|Instituto de Medicina Oncológica y Molecular de Asturias (IMOMA), Oviedo, Spain.
authors.forename: Juan
authors.initials: J
authors.lastname: Cadiñanos
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Quail
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: George S
authors.identifier: http://orcid.org/0000-0003-4337-8022
authors.initials: GS
authors.lastname: Vassiliou
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Pentao
authors.initials: P
authors.lastname: Liu
authors.affiliation: The Wellcome Trust Sanger Institute, Genome Campus, Hinxton/Cambridge, UK.
authors.forename: Allan
authors.initials: A
authors.lastname: Bradley
authors.affiliation: Department of Medicine II, Klinikum rechts der Isar, Technische Universität München, Munich, Germany.|German Cancer Research Center (DKFZ), Heidelberg, &German Cancer Consortium (DKTK), Heidelberg, Germany.
authors.forename: Roland
authors.initials: R
authors.lastname: Rad
mesh_terms: D000818:Animals; D053938:DNA Fragmentation; D004252:DNA Mutational Analysis / Q000379:methods*; D004251:DNA Transposable Elements / Q000235:genetics*; D053263:Gene Regulatory Networks; D023281:Genomics / Q000379:methods*; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D016296:Mutagenesis; D016254:Mutagenesis, Insertional; D009369:Neoplasms / Q000235:genetics*; D009690:Nucleic Acid Conformation
chemical_list: D004251:DNA Transposable Elements