No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice.
CRISPR-Cas9 technologies have transformed genome-editing of experimental organisms and have immense therapeutic potential. Despite significant advances in our understanding of the CRISPR-Cas9 system, concerns remain over the potential for off-target effects. Recent studies have addressed these conce...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2018-07-01
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| Series: | PLoS Genetics |
| Online Access: | https://journals.plos.org/plosgenetics/article/file?id=10.1371/journal.pgen.1007503&type=printable |
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| author | Vivek Iyer Katharina Boroviak Mark Thomas Brendan Doe Laura Riva Edward Ryder David J Adams |
| author_facet | Vivek Iyer Katharina Boroviak Mark Thomas Brendan Doe Laura Riva Edward Ryder David J Adams |
| author_sort | Vivek Iyer |
| collection | DOAJ |
| description | CRISPR-Cas9 technologies have transformed genome-editing of experimental organisms and have immense therapeutic potential. Despite significant advances in our understanding of the CRISPR-Cas9 system, concerns remain over the potential for off-target effects. Recent studies have addressed these concerns using whole-genome sequencing (WGS) of gene-edited embryos or animals to search for de novo mutations (DNMs), which may represent candidate changes introduced by poor editing fidelity. Critically, these studies used strain-matched, but not pedigree-matched controls and thus were unable to reliably distinguish generational or colony-related differences from true DNMs. Here we used a trio design and whole genome sequenced 8 parents and 19 embryos, where 10 of the embryos were mutagenised with well-characterised gRNAs targeting the coat colour Tyrosinase (Tyr) locus. Detailed analyses of these whole genome data allowed us to conclude that if CRISPR mutagenesis were causing SNV or indel off-target mutations in treated embryos, then the number of these mutations is not statistically distinguishable from the background rate of DNMs occurring due to other processes. |
| format | Article |
| id | doaj-art-ca998000b345459b90eea414bd147196 |
| institution | DOAJ |
| issn | 1553-7390 1553-7404 |
| language | English |
| publishDate | 2018-07-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS Genetics |
| spelling | doaj-art-ca998000b345459b90eea414bd1471962025-08-20T02:45:55ZengPublic Library of Science (PLoS)PLoS Genetics1553-73901553-74042018-07-01147e100750310.1371/journal.pgen.1007503No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice.Vivek IyerKatharina BoroviakMark ThomasBrendan DoeLaura RivaEdward RyderDavid J AdamsCRISPR-Cas9 technologies have transformed genome-editing of experimental organisms and have immense therapeutic potential. Despite significant advances in our understanding of the CRISPR-Cas9 system, concerns remain over the potential for off-target effects. Recent studies have addressed these concerns using whole-genome sequencing (WGS) of gene-edited embryos or animals to search for de novo mutations (DNMs), which may represent candidate changes introduced by poor editing fidelity. Critically, these studies used strain-matched, but not pedigree-matched controls and thus were unable to reliably distinguish generational or colony-related differences from true DNMs. Here we used a trio design and whole genome sequenced 8 parents and 19 embryos, where 10 of the embryos were mutagenised with well-characterised gRNAs targeting the coat colour Tyrosinase (Tyr) locus. Detailed analyses of these whole genome data allowed us to conclude that if CRISPR mutagenesis were causing SNV or indel off-target mutations in treated embryos, then the number of these mutations is not statistically distinguishable from the background rate of DNMs occurring due to other processes.https://journals.plos.org/plosgenetics/article/file?id=10.1371/journal.pgen.1007503&type=printable |
| spellingShingle | Vivek Iyer Katharina Boroviak Mark Thomas Brendan Doe Laura Riva Edward Ryder David J Adams No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. PLoS Genetics |
| title | No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. |
| title_full | No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. |
| title_fullStr | No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. |
| title_full_unstemmed | No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. |
| title_short | No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice. |
| title_sort | no unexpected crispr cas9 off target activity revealed by trio sequencing of gene edited mice |
| url | https://journals.plos.org/plosgenetics/article/file?id=10.1371/journal.pgen.1007503&type=printable |
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