Group Accession SAMEG42083

Submission Title:Study_of_the_genomic_damage_caused_transposition_in_mouse_somatic_cells
Submission Identifier:GEN-ERP000618
Submission Description:We recently described a humanised conditional knock-in mouse model of the main form of NPM1c1. In this model, activation of Npm1c mutations in haemopoietic stem cells (HSC) by Mx1-Cre caused, amongst other effects, Hox gene overexpression, myeloid expansion and increased self-renewal of haemopoietic progenitors. Approximately ? of mice developed AML after a long latency (median 17.5 months), suggesting a requirement for cooperating mutations. To identify such mutations, we generated transgenic mice carrying the Sleeping Beauty (SB)2-3 transposon GrOnc, capable of both gene activation and disruption1. Conditional mobilisation of GrOnc in HSCs caused rapid-onset AML (median 99 days) in 80% of Npm1cA/+ mice, whilst lymphoid malignancies predominated in wild-type mutagenised animals. Mutually exclusive activating insertions in Csf2, Flt3, Rasgrp1 or Kras were found in 57 of 70 leukaemias. Recurrent integrations were also identified in several known and putative novel leukaemia genes including Nf1, Dleu2, Nup98 and Bach2. In parallel to this work we have been studying the leukaemogenic effects of the novel highly mutagenic PiggyBac (PB) transposon developed as a cancer gene discovery model in Allan Bradley?s lab4. One critical difference between the two transposons is the fact that SB exhibits a much higher degree of local hopping around its genomic donor site and also generates genomic footprints usually, but not always, in the form of 6 basepair deletions5. PB usually leave a footprint only occasionally (~1% of mobilisation events)6. Add we have generated data that SB may insert proxinmal to cancer gene and comtinue jumping until it lands in a favourable location within the gene locus (wrt oncogenic transformation efficiency) In order to investigate the degree of genomic damage cause around donor loci, on the donor chromosome and in the proximity of cancer gene, we generated an RNA bait set to ?pull-down? the relevant genomic regions and study them by deep sequencing. For this pilot project we will study 10 samples, including 3 from SB tumours, 3 from PB tumours and 1 unmobilised control. The bait set will also pull down the end of transposon repeats in order to map genomic integrations. We wish to multiplex these 10 pull down samples in one lane of HiSEQ. References 1 Vassiliou, G. S. et al. Leukaemia initiation and progression by mutant Nucleophosmin and synergistic pathways. Nature Genetics 2 Collier, L. S., Carlson, C. M., Ravimohan, S., Dupuy, A. J. & Largaespada, D. A. Cancer gene discovery in solid tumours using transposon-based somatic mutagenesis in the mouse. Nature 436, 272-276 3 Dupuy, A. J., Akagi, K., Largaespada, D. A., Copeland, N. G. & Jenkins, N. A. Mammalian mutagenesis using a highly mobile somatic Sleeping Beauty transposon system. Nature 436, 221-226 4 Rad, R. et al. PiggyBac Transposon Mutagenesis: A Tool for Cancer Gene Discovery in Mice. Science 5 Wang, W. et al. Chromosomal transposition of PiggyBac in mouse embryonic stem cells. Proc Natl Acad Sci U S A 105, 9290-9295 6 Yusa, K., Zhou, L., Li, M. A., Bradley, A. & Craig, N. L. A hyperactive piggyBac transposase for mammalian applications. Proc Natl Acad Sci U S A 108, 1531-1536
Submission Release Date:2013-10-07
Submission Update Date:2014-03-02
Submission Reference Layer:false
Databases:ENA SRA Link

URI: ;

ID: ERP000618 ;

Name: The Wellcome Trust Sanger Institute; Address: ; Role: Submitter; Email: ; URI:
Group Name : Other Group
Samples in group:10
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