A Rapid and Cost-Effective Method for Genotyping Genome-Edited Animals: A Heteroduplex Mobility Assay Using Microfluidic Capillary Electrophoresis

Vanessa Chenouard 1, 2, 3 Lucas Brusselle 1, 2, 3 Jean-Marie Heslan 4 Severine Remy 1, 2, 3 Séverine Ménoret 1, 2, 3 Claire Usal 1, 2, 3 Laure-Hélène Ouisse 1, 2, 3 Tuan Nguyen 4 Ignacio Anegon 1, 2, 3 Laurent Tesson 1, 2, 3, *
* Corresponding author
3 P2R - INSERM UMS016/CNRS UMS3556/UN FED4203 - Plateforme "Production de protéines recombinantes"
SFR François Bonamy - Structure fédérative de recherche François Bonamy
4 Inserm/ITUN/CHU Nantes-IFR Santé François Bonamy - plate-forme GenoCellEdit
U1064 Inserm - CRTI - Centre de Recherche en Transplantation et Immunologie, ITUN - Institut de Transplantation Urologie-Néphrologie
Abstract : The recent emergence and application of engineered endonucleases have led to the development of genome editing tools capable of rapidly implementing various targeted genome editions in a wide range of species. Moreover, these novel tools have become easier to use and have resulted in a great increase of applications. Whilst gene knockout (KO) or knockin (KI) animal models are relatively easy to achieve, there is a bottleneck in the detection and analysis of these mutations. Although several methods exist to detect these targeted mutations, we developed a heteroduplex mobility assay on an automated microfluidic capillary electrophoresis system named HMA-CE in order to accelerate the genotyping process. The HMA-CE method uses a simple PCR amplification of genomic DNA (gDNA) followed by an automated capillary electrophoresis step which reveals a heteroduplexes (HD) signature for each mutation. This allows efficient discrimination of wild-type and genome-edited animals down to the single base pair level.
Document type :
Journal articles
Complete list of metadatas

Cited literature [18 references]  Display  Hide  Download

https://www.hal.inserm.fr/inserm-02150677
Contributor : Ana Paula Dutra Azevedo <>
Submitted on : Friday, June 7, 2019 - 1:19:40 PM
Last modification on : Saturday, June 8, 2019 - 1:25:37 AM

File

 Restricted access
To satisfy the distribution rights of the publisher, the document is embargoed until : jamais

Please log in to resquest access to the document

Identifiers

Collections

Citation

Vanessa Chenouard, Lucas Brusselle, Jean-Marie Heslan, Severine Remy, Séverine Ménoret, et al.. A Rapid and Cost-Effective Method for Genotyping Genome-Edited Animals: A Heteroduplex Mobility Assay Using Microfluidic Capillary Electrophoresis. International Journal of Genetics and Genomics, Science Publishing Group, 2016, 43 (5), pp.341-348. ⟨10.1016/j.jgg.2016.04.005⟩. ⟨inserm-02150677⟩

Share

Metrics

Record views

35