A Rapid and Cost-Effective Method for Genotyping Genome-Edited Animals: A Heteroduplex Mobility Assay Using Microfluidic Capillary Electrophoresis. (20th May 2016)

• Record Type: Journal Article
• Authors: Chenouard, Vanessa; Brusselle, Lucas; Heslan, Jean-Marie; Remy, Séverine; Ménoret, Séverine; Usal, Claire; Ouisse, Laure-Hélène; NGuyen, Tuan Huy; Anegon, Ignacio; Tesson, Laurent
• Is Part Of: Journal of genetics and genomics. Volume 43:Number 5(2016)
• Page Start: 341
• Page End: 348
• Publication Date: 2016-05-20
• Subjects: Genome editing -- Genotyping -- Heteroduplex mobility assay -- Capillary electrophoresis; Genetics -- Periodicals; Genomics -- Periodicals; 576.505
• DOI: 10.1016/j.jgg.2016.04.005
• Languages: English
• ISSNs: 1673-8527

Abstract

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.