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...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.展开更多
基金supported by the fundings from the Region Pays de la Loire through Biogenouest,the IBiSA Program,Fondation Progreffethe French Government through the "Investissement d'avenir" program "TEFOR" project,managed by the National Research Agency(No.ANR-II-INSB-0014)the context of the "Investissement d'avenir" program LabEX IGO of the IHU-CESTI projects managed by the National Research Agency(Nos.ANR-11-LABX-001601 and ANR-10-IBHU-005,respectively)
文摘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.
