Project description:Definition using LAM-PCR of the insertion profile of IFP2 transposons containing a NeoR using the wild type CRD-less PB transposase (1-558)
Project description:Definition using LAM-PCR of the insertion profile of IFP2 transposons containing a NeoR using the wild type CRD-less PB transposase (1-558) N-terminally fused with a SV40 NLS
Project description:We have developed a microarray intended for use in finding all transposons in a region of interest. By selectively amplifying and hybridizing transposon flanking DNA to our array, we can localize all transposons in the region present on our TIP-chip, a dense tiling array. We have tested our technology in yeast and have been successful. Keywords: transposon insertion profiling, genomic DNA, yeast
Project description:Genomic rearrangements are a hallmark of childhood cancers, but the mutational processes underlying most of these variants remain unknown. We identified piggyBac transposable element derived 5 (PGBD5) as a highly expressed, enzymatically active domesticated human DNA transposase in a large subset of pediatric solid tumors, including rhabdoid tumors. Ectopic expression of PGBD5 in primary human cells was sufficient to induce fully penetrant cell transformation both in vitro and in immunodeficient mice in vivo. This activity required specific catalytic aspartic acid residues in the PGBD5 transposase domain as well as cellular non-homologous end-joining DNA repair, and was associated with distinct structural rearrangements defined by specific DNA sequence motifs. Similar genomic alterations, some recurrent, were found in primary human rhabdoid tumors. Thus, PGBD5 represents a new class of developmental oncogenic mutators in childhood solid tumors.
Project description:Mammalian chromosomes are partitioned into contact domains that can be conserved as functional units in evolution. Disruptions of domains can result from perturbed CTCF, cohesin, or chromosomal rearrangements. However, to what extent domains can be created de novo has not been explored in depth. Here, using a gain-of-function approach leveraging genome editing and Hi-C, we examined whether, and how, a putative boundary element can function to organize de novo domains in the context of multiple ectopic insertion sites. We subsequently dissected the distinct roles of the CTCF binding site and the transcription start site within the insertion element in changing genome folding.
Project description:Mammalian chromosomes are partitioned into contact domains that can be conserved as functional units in evolution. Disruptions of domains can result from perturbed CTCF, cohesin, or chromosomal rearrangements. However, to what extent domains can be created de novo has not been explored in depth. Here, using a gain-of-function approach leveraging genome editing and Hi-C, we examined whether, and how, a putative boundary element can function to organize de novo domains in the context of multiple ectopic insertion sites. We subsequently dissected the distinct roles of the CTCF binding site and the transcription start site within the insertion element in changing genome folding.