Skip to main content
bioRxiv
  • Home
  • About
  • Submit
  • ALERTS / RSS
Advanced Search
New Results

A marker-free co-selection strategy for high efficiency human genome engineering

Daniel Agudelo, Lusiné Bozoyan, Alexis Duringer, Caroline C. Huard, Sophie Carter, Jeremy Loehr, Dafni Synodinou, Mathieu Drouin, Jayme Salsman, Graham Dellaire, Josée Laganière, View ORCID ProfileYannick Doyon
doi: https://doi.org/10.1101/116251
Daniel Agudelo
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Lusiné Bozoyan
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
2Research and Development, Héma-Québec, Quebec City, QC G1V 5C3, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Alexis Duringer
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Caroline C. Huard
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sophie Carter
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jeremy Loehr
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Dafni Synodinou
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Mathieu Drouin
2Research and Development, Héma-Québec, Quebec City, QC G1V 5C3, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jayme Salsman
3Department of Pathology, Dalhousie University, Halifax, Nova Scotia, B3H 4R2, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Graham Dellaire
3Department of Pathology, Dalhousie University, Halifax, Nova Scotia, B3H 4R2, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Josée Laganière
2Research and Development, Héma-Québec, Quebec City, QC G1V 5C3, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yannick Doyon
1Centre Hospitalier Universitaire de Québec Research Center and Faculty of Medicine, Laval University, Quebec City, QC G1V 4G2, Canada.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Yannick Doyon
  • For correspondence: Yannick.Doyon@crchudequebec.ulaval.ca
  • Abstract
  • Full Text
  • Info/History
  • Metrics
  • Supplementary material
  • Preview PDF
Loading

ABSTRACT

Targeted genome editing using engineered nucleases facilitates the creation of bona fide cellular models for biological research and may be applied to human cell-based therapies. Broadly applicable and versatile methods for increasing the levels of gene editing in cell populations remain highly desirable due to the variable efficiency between distinct genomic loci and cell types. Harnessing the multiplexing capabilities of CRISPR-Cas9 and Cpf1 systems, we designed a simple and robust co-selection strategy for enriching cells harboring either nuclease-driven non-homologous end joining (NHEJ) or homology-directed repair (HDR) events. Selection for dominant alleles of the endogenous sodium-potassium pump (Na+,K+-ATPase) that render cells resistant to ouabain is used to enrich for custom modifications at another unlinked locus of interest, effectively increasing the recovery of engineered cells. The process was readily adaptable to transformed and primary cells, including hematopoietic stem and progenitor cells (HSPCs). The use of universal CRISPR reagents and a commercially available small molecule inhibitor streamlines the incorporation of marker-free genetic changes in human cells.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
Back to top
PreviousNext
Posted March 15, 2017.
Download PDF

Supplementary Material

Email

Thank you for your interest in spreading the word about bioRxiv.

NOTE: Your email address is requested solely to identify you as the sender of this article.

Enter multiple addresses on separate lines or separate them with commas.
A marker-free co-selection strategy for high efficiency human genome engineering
(Your Name) has forwarded a page to you from bioRxiv
(Your Name) thought you would like to see this page from the bioRxiv website.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
A marker-free co-selection strategy for high efficiency human genome engineering
Daniel Agudelo, Lusiné Bozoyan, Alexis Duringer, Caroline C. Huard, Sophie Carter, Jeremy Loehr, Dafni Synodinou, Mathieu Drouin, Jayme Salsman, Graham Dellaire, Josée Laganière, Yannick Doyon
bioRxiv 116251; doi: https://doi.org/10.1101/116251
Reddit logo Twitter logo Facebook logo LinkedIn logo Mendeley logo
Citation Tools
A marker-free co-selection strategy for high efficiency human genome engineering
Daniel Agudelo, Lusiné Bozoyan, Alexis Duringer, Caroline C. Huard, Sophie Carter, Jeremy Loehr, Dafni Synodinou, Mathieu Drouin, Jayme Salsman, Graham Dellaire, Josée Laganière, Yannick Doyon
bioRxiv 116251; doi: https://doi.org/10.1101/116251

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Subject Area

  • Molecular Biology
Subject Areas
All Articles
  • Animal Behavior and Cognition (4239)
  • Biochemistry (9171)
  • Bioengineering (6804)
  • Bioinformatics (24062)
  • Biophysics (12154)
  • Cancer Biology (9564)
  • Cell Biology (13825)
  • Clinical Trials (138)
  • Developmental Biology (7656)
  • Ecology (11736)
  • Epidemiology (2066)
  • Evolutionary Biology (15540)
  • Genetics (10670)
  • Genomics (14358)
  • Immunology (9511)
  • Microbiology (22901)
  • Molecular Biology (9129)
  • Neuroscience (49112)
  • Paleontology (357)
  • Pathology (1487)
  • Pharmacology and Toxicology (2583)
  • Physiology (3851)
  • Plant Biology (8351)
  • Scientific Communication and Education (1473)
  • Synthetic Biology (2301)
  • Systems Biology (6205)
  • Zoology (1302)