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Forward and Reverse Genetic Approaches to Studying Ciliopathy in Zebrafish

Khosravi, Maryam; (2019) Forward and Reverse Genetic Approaches to Studying Ciliopathy in Zebrafish. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Genetic kidney diseases can often present with advanced renal failure, so remains undiagnosed, because urgent treatment with renal replacement therapy is prioritised over obtaining a genetic diagnosis. // In a population with un-diagnosed familial kidney disease, two families were identified with distinct clinical phenotypes. Two novel mutations in HACL1 and COL4A1 were identified in these patients and their respective connection to cilia and collagen biology was sought. The mutations were modelled using the reverse genetic approach of CRISPR-Cas9 gene editing in zebrafish, and the resultant animal models were studied. After successfully generating zebrafish mutant alleles for the hacl1 disease gene, I went on to study the phenotype and assay the cilia in different organs of the zebrafish. I did not detect an overt phenotype. The desired col4a1 was obtained in injected embryos, but this mutation was not transmitted to further generations. // Using a second approach of forward genetics, I characterised a novel mutant zebrafish line containing a mutation that disrupts function of the Ift144 ciliary protein. This mutant presents with a cystic pronephros and abnormal ciliary morphology. Having characterised this mutant and identified a mutation in ciliary protein ift144 by RNA sequencing, I went on to use this model for drug testing with selected drugs known to rescue renal cysts in other animal models. These drugs all significantly reduced cyst size in the zebrafish mutant, proving the principle that this model has good potential for high throughput in vivo screening of drugs to treat ciliopathies.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Forward and Reverse Genetic Approaches to Studying Ciliopathy in Zebrafish
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Copyright © The Author 2019. Original content in this thesis is licensed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) Licence (https://creativecommons.org/licenses/by/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
UCL classification: UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences
URI: https://discovery.ucl.ac.uk/id/eprint/10079759
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