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Highly specific genome editing in human cells and plants using CRISPR systems

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Authors
김정은
Advisor
이형호
Major
자연과학대학 화학부
Issue Date
2017-02
Publisher
서울대학교 대학원
Keywords
Genome engineeringProgrammable nucleasesClustered Regularly Interspaced Short Palindromic Repeats (CRISPR)RNA-guided Endonucleases (RGENs)Cpf1
Description
학위논문 (박사)-- 서울대학교 대학원 : 화학부, 2017. 2. 이형호.
Abstract
Targeted genome engineering with RNA-guided engineered nucleases (RGENs) repurposed from the clustered regularly interspaced short palindromic repeat (CRISPR) systems has recently received significant attention because of their simple design and preparation. Although, RGENs-mediated genome editing has been successfully performed in cultured cells and model organisms, developing a highly efficient and specific CRISPR-based genome editing platform is still a major challenge in this area.
In this study, I develop and optimize the new CRISPR-based genome editing platforms and investigate their target specificities. First, I present delivery of RGEN RNPs in plant cells. RGEN RNPs enabled DNA-free genome editing in plant cells at high frequencies up to 44%. Whole plants were successfully regenerated from gene-edited cells and no apparent off-target mutations were detected in these mutants. This method may reduce the likelihood of DNA insertion and potential off-target effects and alleviate the concerns related to genetically modified plants. Second, I optimize the Cpf1-mediated genome editing system and investigate its target specificity. CRISPR-Cpf1 system induced targeted genome modification efficiently in human cells. Mismatched crRNAs test and genome-wide off-target analysis showed that this nuclease yielded DNA cleavages at the target site in a highly specific manner. These results indicate that CRISPR-Cpf1 is suitable for precision genome editing and will be a good option for RNA-guided genome editing.
Language
English
URI
https://hdl.handle.net/10371/125337
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College of Natural Sciences (자연과학대학)Dept. of Chemistry (화학부)Theses (Ph.D. / Sc.D._화학부)
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