Transposon mediated integration of plasmid DNA into the subventricular zone of neonatal mice to generate novel models of glioblastoma.
An urgent need exists to test the contribution of new genes to the pathogenesis and progression of human glioblastomas (GBM), the most common primary brain tumor in adults with dismal prognosis. New potential therapies are rapidly emerging from the bench and require systematic testing in experimental models which closely reproduce the salient features of the human disease. Herein we describe in detail a method to induce new models of GBM with transposon-mediated integration of plasmid DNA into cells of the subventricular zone of neonatal mice. We present a simple way to clone new transposons amenable for genomic integration using the Sleeping Beauty transposon system and illustrate how to monitor plasmid uptake and disease progression using bioluminescence, histology and immuno-histochemistry. We also describe a method to create new primary GBM cell lines. Ideally, this report will allow further dissemination of the Sleeping Beauty transposon system among brain tumor researchers, leading to an in depth understanding of GBM pathogenesis and progression and to the timely design and testing of effective therapies for patients.
Department of Neurosurgery, University of Michigan School of Medicine.
Anda-Alexandra
AA
Calinescu
Department of Neurosurgery, University of Michigan School of Medicine.
Felipe Javier
FJ
Núñez
Department of Pediatrics, Division of Hematology-Oncology, University of Michigan School of Medicine.
Carl
C
Koschmann
Department of Neurosurgery, University of Michigan School of Medicine.
Bradley L
BL
Kolb
Department of Neurosurgery, University of Michigan School of Medicine; Department of Cell and Developmental Biology, University of Michigan.
Pedro R
PR
Lowenstein
Department of Neurosurgery, University of Michigan School of Medicine; Department of Cell and Developmental Biology, University of Michigan; mariacas@umich.edu.
Maria G
MG
Castro
D000818:Animals; D001932:Brain Neoplasms / Q000235:genetics*; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D004251:DNA Transposable Elements; D004195:Disease Models, Animal; D005260:Female; D005909:Glioblastoma / Q000235:genetics*; D006801:Humans; D007276:Injections, Intraventricular; D020547:Lateral Ventricles / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D011247:Pregnancy; D019895:Transposases / Q000235:genetics / Q000378:metabolism
D004251:DNA Transposable Elements; D004247:DNA; D019895:Transposases