Genome-wide transposon screening and quantitative insertion site sequencing for cancer gene discovery in mice

Mathias J. Friedrich, Lena Rad, Iraad F. Bronner, Alexander Strong, Wei Wang, Julia Weber, Matthew Mayho, Hannes Ponstingl, Thomas Engleitner, Carolyn Grove, Anja Pfaus, Dieter Saur, Juan Cadiñanos, Michael A. Quail, George S. Vassiliou, Pentao Liu, Allan Bradley, Roland Rad

Publikation: Beitrag in FachzeitschriftArtikelBegutachtung

38 Zitate (Scopus)


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.

Seiten (von - bis)289-309
FachzeitschriftNature Protocols
PublikationsstatusVeröffentlicht - 1 Feb. 2017
Extern publiziertJa


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