Date of Completion


Embargo Period



Anastasios Tzingounis, Daniel Mulkey

Field of Study

Physiology and Neurobiology


Master of Science

Open Access

Open Access


Neurofibramatosis type 1 is an autosomal dominant disorder characterized by the development of both benign and malignant neoplasms of glial origin, as well as learning deficits and, less frequently, macrocephaly. Essential to the understanding of NF1 is the observation that the peripheral neoplasms (neurofibroma) which develop are composed primarily of NF1 negative glia which have undergone a loss of NF1 heterozygosity. The etiology of this loss of heterozygosity remains unclear, but a better characterization of these mutation events would provide insight into the cellular origins of the NF1 phenotype. CRISPR/Cas9 systems for genome editing have been, since 2013, a significant technique of interest by which precise mutagenesis may be achieved. Here we demonstrate a use of this system in conjunction with a Piggybac transposon lineage labelling approach by which the clonal identity of modified cell populations and the influence on cortical phenotype of a CRISPR/Cas9 induced model of NF1 deficiency may be investigated. Characterization of the induced NF1 phenotype is performed, and the CRISPR/Cas9 system is shown to be an effective method by which loss of cellular heterozygosity by somatic mutation may be modelled, exhibiting a monoallelic mutation rate of between 42.2% and 48.5. NF1 condition glia are not found to exhibit abnormal proliferation in the adult brain, and evidence supporting a model of neural progenitor fate switching to a glial phenotype in the form of CAG-TDTomato positive astrocyte populations derived from neuronal precursors is presented.

Major Advisor

Joseph Loturco