Direct production of mouse disease models by embryo microinjection of TALENs and oligodeoxynucleotides

The study of genetic disease mechanisms relies mostly on targeted mouse mutants that are derived from engineered embryonic stem (ES) cells. Nevertheless, the establishment of mutant ES cells is laborious and time-consuming, restricting the study of the increasing number of human disease mutations di...

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Veröffentlicht in:Proceedings of the National Academy of Sciences - PNAS 2013-03, Vol.110 (10), p.3782-3787
Hauptverfasser: Wefers, Benedikt, Meyer, Melanie, Ortiz, Oskar, de Angelis, Martin Hrabé, Hansen, Jens, Wurst, Wolfgang, Kühn, Ralf
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container_end_page 3787
container_issue 10
container_start_page 3782
container_title Proceedings of the National Academy of Sciences - PNAS
container_volume 110
creator Wefers, Benedikt
Meyer, Melanie
Ortiz, Oskar
de Angelis, Martin Hrabé
Hansen, Jens
Wurst, Wolfgang
Kühn, Ralf
description The study of genetic disease mechanisms relies mostly on targeted mouse mutants that are derived from engineered embryonic stem (ES) cells. Nevertheless, the establishment of mutant ES cells is laborious and time-consuming, restricting the study of the increasing number of human disease mutations discovered by high-throughput genomic analysis. Here, we present an advanced approach for the production of mouse disease models by microinjection of transcription activator-like effector nucleases (TALENs) and synthetic oligodeoxynucleotides into one-cell embryos. Within 2 d of embryo injection, we created and corrected chocolate missense mutations in the small GTPase RAB38; a regulator of intracellular vesicle trafficking and phenotypic model of Hermansky-Pudlak syndrome. Because ES cell cultures and targeting vectors are not required, this technology enables instant germline modifications, making heterozygous mutants available within 18 wk. The key features of direct mutagenesis by TALENs and oligodeoxynucleotides, minimal effort and high speed, catalyze the generation of future in vivo models for the study of human disease mechanisms and interventions.
doi_str_mv 10.1073/pnas.1218721110
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Nevertheless, the establishment of mutant ES cells is laborious and time-consuming, restricting the study of the increasing number of human disease mutations discovered by high-throughput genomic analysis. Here, we present an advanced approach for the production of mouse disease models by microinjection of transcription activator-like effector nucleases (TALENs) and synthetic oligodeoxynucleotides into one-cell embryos. Within 2 d of embryo injection, we created and corrected chocolate missense mutations in the small GTPase RAB38; a regulator of intracellular vesicle trafficking and phenotypic model of Hermansky-Pudlak syndrome. Because ES cell cultures and targeting vectors are not required, this technology enables instant germline modifications, making heterozygous mutants available within 18 wk. 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subjects Alleles
Amino Acid Sequence
Animals
Base Sequence
Biological Sciences
Cleavage Stage, Ovum
Deoxyribonucleases - administration & dosage
Deoxyribonucleases - genetics
Disease models
Disease Models, Animal
DNA
Embryos
Exons
Female
Genetic Diseases, Inborn - genetics
Genetic disorders
Genetic mutation
Genetic Techniques
Genetic Vectors
Genomes
Genomics
Genotype & phenotype
Germ-Line Mutation
HEK293 Cells
Humans
Male
Mice
Mice, Transgenic
Microinjections
Molecular Sequence Data
Mutagenesis
Mutation
Mutation, Missense
Oligodeoxyribonucleotides - administration & dosage
Oligodeoxyribonucleotides - genetics
Polymerase chain reaction
Pregnancy
Pups
rab GTP-Binding Proteins - genetics
Rodents
Sequence Homology, Nucleic Acid
Stem cells
title Direct production of mouse disease models by embryo microinjection of TALENs and oligodeoxynucleotides
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