Current Protocols in Stem Cell BiologyVolume 44, Issue 1 p. 5B.7.1-5B.7.22 UNIT CRISPR-Cas9-Based Genome Editing of Human Induced Pluripotent Stem Cells Joseph C. Giacalone, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, Iowa These authors contributed equally to this manuscriptSearch for more papers by this authorTasneem P. Sharma, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, Iowa These authors contributed equally to this manuscriptSearch for more papers by this authorErin R. Burnight, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorJohn F. Fingert, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorRobert F. Mullins, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorEdwin M. Stone, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorBudd A. Tucker, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this author Joseph C. Giacalone, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, Iowa These authors contributed equally to this manuscriptSearch for more papers by this authorTasneem P. Sharma, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, Iowa These authors contributed equally to this manuscriptSearch for more papers by this authorErin R. Burnight, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorJohn F. Fingert, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorRobert F. Mullins, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorEdwin M. Stone, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this authorBudd A. Tucker, Stephen A. Wynn Institute for Vision Research, Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IowaSearch for more papers by this author Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URLShare a linkShare onEmailFacebookTwitterLinked InRedditWechat Abstract Human induced pluripotent stem cells (hiPSCs) are the ideal cell source for autologous cell replacement. However, for patients with Mendelian diseases, genetic correction of the original disease-causing mutation is likely required prior to cellular differentiation and transplantation. The emergence of the CRISPR-Cas9 system has revolutionized the field of genome editing. By introducing inexpensive reagents that are relatively straightforward to design and validate, it is now possible to correct genetic variants or insert desired sequences at any location within the genome. CRISPR-based genome editing of patient-specific iPSCs shows great promise for future autologous cell replacement therapies. One caveat, however, is that hiPSCs are notoriously difficult to transfect, and optimized experimental design considerations are often necessary. This unit describes design strategies and methods for efficient CRISPR-based genome editing of patient- specific iPSCs. Additionally, it details a flexible approach that utilizes positive selection to generate clones with a desired genomic modification, Cre-lox recombination to remove the integrated selection cassette, and negative selectionto eliminate residual hiPSCs with intact selection cassettes. © 2018 by John Wiley & Sons, Inc. Please check your email for instructions on resetting your password. If you do not receive an email within 10 minutes, your email address may not be registered, and you may need to create a new Wiley Online Library account. Can\'t sign in? Forgot your username? Enter your email address below and we will send you your username If the address matches an existing account you will receive an email with instructions to retrieve your username