Forward genetic insertional mutagenesis screens are used by many labs to identify candidate cancer genes. We and others have used the Sleeping Beauty DNA transposon to generate random mutations within the murine genome that cause cancer. Identification of the insertion sites, either via RNA sequencing or DNA sequencing, is required for cancer gene discovery. Multiple sequencing-based approaches have been utilized to identify locations of transposon insertions within a genome including linker-mediated PCR, RNA-Seq, and Seq capture. Here, a bioinformatics pipeline is presented applicable to both the identification of transposon-generated fusions in RNA-Seq data and the direct identification of transposon insertion sites in DNA sequencing data. We are currently utilizing this method to identify transposon insertions generated by Sleeping Beauty transposase-mediated mobilization of the T2/Onc transposon within the murine genome. With slight modification, this approach is amenable to the identification of any mobile genetic element within any genome.
|Original language||English (US)|
|Title of host publication||Methods in Molecular Biology|
|Publisher||Humana Press Inc.|
|Number of pages||7|
|State||Published - 2019|
|Name||Methods in Molecular Biology|
Bibliographical noteFunding Information:
This work was supported by NCI grant R50-CA211249 to Aaron Sarver.
- Insertion site