Genome editing approach for generation of isogenic cell lines modelling Huntington's disease in vitro
- Authors: Malakhova A.A1,2,3, Sorokin M.A1,2,3, Sorokina A.E1,2,3, Malankhanova T.B1,2,3,4, Mazurok N.A1,2,3, Medvedev S.P1,2,3, Zakian S.M1,2,3
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Affiliations:
- Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences
- Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences
- State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian Federation
- National Research University Novosibirsk State University
- Issue: Vol 11, No 2 (2016)
- Pages: 106-113
- Section: Articles
- URL: https://genescells.ru/2313-1829/article/view/120601
- DOI: https://doi.org/10.23868/gc120601
- ID: 120601
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Abstract
Huntington's disease is an autosomal dominant neurodegenerative disorder caused by trinucleotide repeats expansion mutation in the first exon of HTT gene. Neurodegenerative disorders are difficult to study due to limited material availability and late onset of the manifestation. In vitro cell models provide a good opportunity to study molecular mechanisms of the pathology. New genome editing approaches allow us to generate isogenic cell lines that can be useful for drug screening and studying mechanisms of molecular and cellular events triggered by certain mutation on an equal genetic background. The aim of our work was establishing a method to introduce expanded CAG repeat tract into HTT gene by homology directed repair. We used a genome editing tool based on CRISPR/Cas9 system to facilitate the process. A donor construct bearing 215 CAG trinucleotide repeats flanked with long homology arms was generated. The method was approved on the HEK293Phoenix cell line. Mutant clones were obtained. It was shown that mutant HTT protein is expressed in the HEK293 mutant clones and affect their viability and proliferation rate.
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About the authors
A. A Malakhova
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian Federation
Email: amal@bionet.nsc.ru
Novosibirsk, Russia
M. A Sorokin
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian FederationNovosibirsk, Russia
A. E Sorokina
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian FederationNovosibirsk, Russia
T. B Malankhanova
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian Federation; National Research University Novosibirsk State UniversityNovosibirsk, Russia
N. A Mazurok
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian FederationNovosibirsk, Russia
S. P Medvedev
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian FederationNovosibirsk, Russia
S. M Zakian
Federal Research Center Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences; Institute of Chemical Biology and Fundamental Medicine, the Siberian Branch of the Russian Academy of Sciences; State Research Institute of Circulation Pathology, Ministry of Healthcare of the Russian FederationNovosibirsk, Russia
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