3.4 Genome Editing with CRISPR/Cas9 v1

Abstract

This is part 3.4 of the "Induced Neurons for the Study of Neurodegenerative and Neurodevelopmental Disorders" collection of protocols. Collection Abstract: Patient-derived or genomically modified human induced pluripotent stem cell s (iPSCs) offer the opportunity to study neurodevelopmental and neurodegenerative disorders. Overexpression of certain neurogenic transcription factors (TFs) in iPSCs can induce efficient differentiation into homogeneous populations of the disease-relevant neuronal cell types. Here we provide protocols for genomic manipulations of iPSCs by CRISPR/Cas9. We also introduce two methods, based on lentiviral delivery and the piggyBac transposon system, to stably integrate neurogenic TFs into human iPSCs. Furthermore, we describe the TF-mediated neuronal differentiation and maturation in combination with astrocyte cocultures.