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A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh

The introduction of CRISPR-Cas9 technology for targeted mutagenesis has revolutionized reverse genetics and made genome editing a realistic option in many model organisms. One of the difficulties with this technique is screening for mutations in large numbers of samples. Many screening approaches fo...

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Autores principales: VanLeuven, Ariel J, Park, Sungdae, Menke, Douglas B, Lauderdale, James D
Formato: Online Artículo Texto
Lenguaje:English
Publicado: 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9580339/
https://www.ncbi.nlm.nih.gov/pubmed/29939088
http://dx.doi.org/10.2144/btn-2018-0012
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author VanLeuven, Ariel J
Park, Sungdae
Menke, Douglas B
Lauderdale, James D
author_facet VanLeuven, Ariel J
Park, Sungdae
Menke, Douglas B
Lauderdale, James D
author_sort VanLeuven, Ariel J
collection PubMed
description The introduction of CRISPR-Cas9 technology for targeted mutagenesis has revolutionized reverse genetics and made genome editing a realistic option in many model organisms. One of the difficulties with this technique is screening for mutations in large numbers of samples. Many screening approaches for identifying CRISPR-Cas9 mutants have been published; however, in practice these methods are time consuming, expensive, or often yield false positives. This report describes a PCR-based screening approach using non-denaturing PAGE. This approach does not depend on the formation of heteroduplexes and reliably detects changes as small as 1 base-pair (bp) in nucleic acid length at the target site. This approach can be used to identify novel mutations and is also useful as a routine genotyping method.
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spelling pubmed-95803392022-10-19 A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh VanLeuven, Ariel J Park, Sungdae Menke, Douglas B Lauderdale, James D Biotechniques Article The introduction of CRISPR-Cas9 technology for targeted mutagenesis has revolutionized reverse genetics and made genome editing a realistic option in many model organisms. One of the difficulties with this technique is screening for mutations in large numbers of samples. Many screening approaches for identifying CRISPR-Cas9 mutants have been published; however, in practice these methods are time consuming, expensive, or often yield false positives. This report describes a PCR-based screening approach using non-denaturing PAGE. This approach does not depend on the formation of heteroduplexes and reliably detects changes as small as 1 base-pair (bp) in nucleic acid length at the target site. This approach can be used to identify novel mutations and is also useful as a routine genotyping method. 2018-06 /pmc/articles/PMC9580339/ /pubmed/29939088 http://dx.doi.org/10.2144/btn-2018-0012 Text en https://creativecommons.org/licenses/by/4.0/This work is licensed under the Creative Commons Attribution 4.0 License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/)
spellingShingle Article
VanLeuven, Ariel J
Park, Sungdae
Menke, Douglas B
Lauderdale, James D
A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title_full A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title_fullStr A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title_full_unstemmed A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title_short A PAGE screening approach for identifying CRISPR-Cas9-induced mutations in zebrafi sh
title_sort page screening approach for identifying crispr-cas9-induced mutations in zebrafi sh
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9580339/
https://www.ncbi.nlm.nih.gov/pubmed/29939088
http://dx.doi.org/10.2144/btn-2018-0012
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