Abstract
Although whole-genome sequencing has uncovered a large number of mutations that drive tumorigenesis, functional ratification for most mutations remains sparse. Here, we present an approach to test functional relevance of tumor mutations employing CRISPR/Cas9. Combining comprehensive sgRNA design and an efficient reporter assay to nominate efficient and selective sgRNAs, we establish a pipeline to dissect roles of cancer mutations with potential applicability to personalized medicine and future therapeutic use.
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MeSH terms
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Bacterial Proteins* / genetics
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Bacterial Proteins* / metabolism
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CRISPR-Associated Protein 9
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Carcinoma / genetics*
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Clustered Regularly Interspaced Short Palindromic Repeats*
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Colonic Neoplasms / genetics*
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Computational Biology
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DNA Cleavage
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Endonucleases* / genetics
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Endonucleases* / metabolism
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Genes, Reporter
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Green Fluorescent Proteins / genetics
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HeLa Cells
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Humans
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Leukemia, Myeloid, Acute / genetics*
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Mutation / genetics*
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Nuclear Proteins / genetics
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Nucleophosmin
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Proto-Oncogene Proteins B-raf / genetics
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RNA, Guide, CRISPR-Cas Systems / genetics*
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Transfection
Substances
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Bacterial Proteins
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Nuclear Proteins
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RNA, Guide, CRISPR-Cas Systems
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Nucleophosmin
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Green Fluorescent Proteins
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BRAF protein, human
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Proto-Oncogene Proteins B-raf
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CRISPR-Associated Protein 9
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Cas9 endonuclease Streptococcus pyogenes
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Endonucleases