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Translocation and duplication from CRISPR-Cas9 editing in Arabidopsis thaliana

Peter G. Lynagh, Soichi Inagaki, Kirk R. Amundson, Mohan P.A. Marimuthu, Brett Randolph Pike, Isabelle M. Henry, Ek Han Tan, View ORCID ProfileLuca Comai
doi: https://doi.org/10.1101/400507
Peter G. Lynagh
UC Davis Genome Center and Department of Plant Biology, Davis, California
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Soichi Inagaki
UC Davis Genome Center and Department of Plant Biology, Davis, CaliforniaNational Institute of Genetics, Mishima, Japan
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Kirk R. Amundson
UC Davis Genome Center and Department of Plant Biology, Davis, California
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Mohan P.A. Marimuthu
UC Davis Genome Center and Department of Plant Biology, Davis, California
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Brett Randolph Pike
UC Davis Genome Center and Department of Plant Biology, Davis, California
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Isabelle M. Henry
UC Davis Genome Center and Department of Plant Biology, Davis, California
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Ek Han Tan
UC Davis Genome Center and Department of Plant Biology, Davis, CaliforniaSchool of Biology and Ecology, University of Maine, Orono, Maine
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Luca Comai
UC Davis Genome Center and Department of Plant Biology, Davis, California
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  • ORCID record for Luca Comai
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Abstract

Cut DNA ends in plants may recombine to form novel molecules. We asked whether CRISPR-Cas9 expression in plants could induce nonhomologous recombination between diverse and heterologous broken DNA ends. We induced two breaks separated by 2.3 or by 8.5 kilobases leading to duplication of the intervening DNA and meiotic transmission of the 2.3kb duplication. Two or more dsDNA breaks in nonhomologous chromosomes led to ligation of breakpoints consistent with chromosome arm translocations. Screening 881 primary transformants we obtained 195 PCR products spanning independent, expected translocation junctions involving ends produced by cutting different loci. Sequencing indicated a true positive rate of 84/91 and demonstrated the occurrence of different junction alleles. A majority of the resulting structures would be deleterious and none were transmitted meiotically. Ligation of interchromosomal, heterologous dsDNA ends suggest that the CRISPR-Cas9 can be used to engineer plant genes and chromosomes in vivo.

Significance Statement We explored how genome editing tools such as CRISPR-Cas9 could provide new ways to tailor novel genomic combinations and arrangements. We show that distant cut ends often precisely come together, that cuts in different chromosomes can result in translocations, and that two cuts within a chromosome often result in the duplication of the intervening segment. Formation of multiple structures with precise junctions will enable engineered rearrangements that can be predicted with accuracy.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted August 26, 2018.
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Translocation and duplication from CRISPR-Cas9 editing in Arabidopsis thaliana
Peter G. Lynagh, Soichi Inagaki, Kirk R. Amundson, Mohan P.A. Marimuthu, Brett Randolph Pike, Isabelle M. Henry, Ek Han Tan, Luca Comai
bioRxiv 400507; doi: https://doi.org/10.1101/400507
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Translocation and duplication from CRISPR-Cas9 editing in Arabidopsis thaliana
Peter G. Lynagh, Soichi Inagaki, Kirk R. Amundson, Mohan P.A. Marimuthu, Brett Randolph Pike, Isabelle M. Henry, Ek Han Tan, Luca Comai
bioRxiv 400507; doi: https://doi.org/10.1101/400507

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