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Modeling and correction of structural variations in patient-derived iPSCs using CRISPR/Cas9

Authors
 Chul-Yong Park  ;  Jin Jea Sung  ;  Sang-Hwi Choi  ;  Dongjin R Lee  ;  In-Hyun Park  ;  Dong-Wook Kim 
Citation
 NATURE PROTOCOLS, Vol.11(11) : 2154-2169, 2016 
Journal Title
NATURE PROTOCOLS
ISSN
 1754-2189 
Issue Date
2016
MeSH
Base Sequence ; CRISPR-Cas Systems/genetics* ; DNA End-Joining Repair/genetics ; Gene Rearrangement/genetics ; Genetic Engineering/methods* ; Genetic Variation* ; Genome, Human/genetics ; Humans ; Induced Pluripotent Stem Cells/metabolism* ; Models, Genetic* ; Repetitive Sequences, Nucleic Acid/genetics
Abstract
Genome engineering technology using engineered nucleases has been rapidly developing, enabling the efficient correction of simple mutations. However, the precise correction of structural variations (SVs) such as large inversions remains limited. Here we describe a detailed procedure for the modeling or correction of large chromosomal rearrangements and short nucleotide repeat expansions using engineered nucleases in human induced pluripotent stem cells (hiPSCs) from a healthy donor and patients with SVs. This protocol includes the delivery of engineered nucleases with no donor template to hiPSCs, and genotyping and derivation/characterization of gene-manipulated hiPSC clones. With engineered nucleases, genomic inversions, reversions, and deletions of short nucleotide expansions can be identified in 2 weeks, and desired clones can be generated in as little as 3-4 weeks. This protocol enables the correction of large inverted segments and short nucleotide repeat expansions in diseases such as hemophilia A, fragile X syndrome, Hunter syndrome, and Friedreich's ataxia.
Full Text
http://www.nature.com/nprot/journal/v11/n11/full/nprot.2016.129.html
DOI
10.1038/nprot.2016.129
Appears in Collections:
1. College of Medicine (의과대학) > Dept. of Physiology (생리학교실) > 1. Journal Papers
Yonsei Authors
Kim, Dong Wook(김동욱) ORCID logo https://orcid.org/0000-0002-5025-1532
Park, Chul Yong(박철용) ORCID logo https://orcid.org/0000-0002-4467-9268
URI
https://ir.ymlib.yonsei.ac.kr/handle/22282913/152501
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