title: GRK5-Knockout Mice Generated by TALEN-Mediated Gene Targeting.
abstract: Transcription activator-like effector nucleases (TALENs) are a new type of engineered nuclease that is very effective for directed gene disruption in any genome sequence. We investigated the generation of mice with genetic knockout (KO) of the G protein-coupled receptor kinase (GRK) 5 gene by microinjection of TALEN mRNA. TALEN vectors were designed to target exons 1, 3, and 5 of the mouse GRK5 gene. Flow cytometry showed that the activity of the TALEN mRNAs targeted to exons 1, 3, and 5 was 8.7%, 9.7%, and 12.7%, respectively. The TALEN mRNA for exon 5 was injected into the cytoplasm of 180 one-cell embryos. Of the 53 newborns, three (5.6%) were mutant founders (F0) with mutations. Two clones from F028 showed a 45-bp deletion and F039 showed the same biallelic non-frame-shifting 3-bp deletions. Three clones from F041 were shown to possess a combination of frame-shifting 2-bp deletions. All of the mutations were transmitted through the germline but not to all progenies (37.5%, 37.5%, and 57.1% for the F028, F039, and F041 lines, respectively). The homozygote GRK5-KO mice for 28 and 41 lines created on F3 progenies and the homozygous genotype was confirmed by PCR, T7E1 assay and sequencing.
authors: a Animal Biotechnology, Graduate School of Future Convergence Technology , Institute of Genetic Engineering, Hankyong National University , Anseong , Republic of Korea.
authors: Tsevelmaa
authors: T
authors: Nanjidsuren
authors: a Animal Biotechnology, Graduate School of Future Convergence Technology , Institute of Genetic Engineering, Hankyong National University , Anseong , Republic of Korea.
authors: Chae-Won
authors: CW
authors: Park
authors: b National Research Center , Korea Research Institute of Bioscience and Biotechnology , Ochang , Republic of Korea.
authors: Bo-Woong
authors: BW
authors: Sim
authors: b National Research Center , Korea Research Institute of Bioscience and Biotechnology , Ochang , Republic of Korea.
authors: Sun-Uk
authors: SU
authors: Kim
authors: b National Research Center , Korea Research Institute of Bioscience and Biotechnology , Ochang , Republic of Korea.
authors: Kyu-Tae
authors: KT
authors: Chang
authors: c Department of Food and Nutrition , Hoseo University , Asan , Republic of Korea.
authors: Myung-Hwa
authors: MH
authors: Kang
authors: a Animal Biotechnology, Graduate School of Future Convergence Technology , Institute of Genetic Engineering, Hankyong National University , Anseong , Republic of Korea.
authors: Kwan-Sik
authors: KS
authors: Min
mesh_terms: D000818:Animals; D005260:Female; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000235:genetics*; D018390:Gene Targeting / Q000379:methods*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout / Q000235:genetics*; D009154:Mutation; D000069896:Transcription Activator-Like Effector Nucleases / Q000235:genetics*
chemical_list: D054774:G-Protein-Coupled Receptor Kinase 5; C517895:Grk5 protein, mouse; D000069896:Transcription Activator-Like Effector Nucleases
keywords: GRK-5; Gene targeting; Knockout; TALEN