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

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.