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Thesis
Investigating the role of miR-9 in retinal development and adult regeneration in zebrafish
University of West Florida Libraries
Bachelor of Science (BS), University of West Florida
2023
Abstract
Retinal disease in humans causes damage to rod and cone photoreceptors, leading to vision loss and potentially blindness. The retina is a central nervous system tissue that contains many specialized cells for sight. Photoreceptors are a critical part of the retina that allow for vision and once these cells are damaged in humans, they cannot be repaired or replaced. However, the model organism, zebrafish (Danio rerio), can regenerate fully functioning retinal photoreceptors in only 14 days. MicroRNAs (miRNAs), short, noncoding RNA sequences, have important roles in retinal regeneration and development, with important controls of gene expression to affect functions in photoreceptor genesis. To better understand the role of a specific miRNA, miR-9, this work investigated miR-9 in both the developing embryonic retina and the adult regenerating retina in zebrafish. The main objective was to quantify miR-9 molecules during retinal neurogenesis, to test the overall hypothesis that miR-9 regulates photoreceptor genesis in the zebrafish retina. Spatially and temporally, the expression of miR-9 was analyzed in the developing and regenerating adult retina using in situ hybridization (ISH) and immunolabeling techniques. Additionally, reverse-transcription followed with quantitative polymerase chain reaction (RT-qPCR) was used to quantify expression of the seven precursor miR-9 molecules and mature miR-9 in adults. Next, miR-9 was investigated using knockdown techniques in the retina of developing embryos and in adult regenerating retina using morpholino oligonucleotides (MO) and CRISPR/Cas9 gene editing techniques. In proliferating cells of the developing retina, miR-9 was expressed at 48 hours post-fertilization (hpf). Knockdown of miR-9 in embryos resulted in a lower number of proliferating cells at 48 hpf and fewer mature photoreceptors at 72 hpf, suggesting miR-9 is important for normal cell proliferation and normal photoreceptor development. In adults subjected to bright-light-induced retinal lesioning, knockdown of miR-9 resulted in a higher number of proliferating cells in the outer nuclear layer (ONL) at 3 dpl, suggesting miR-9 may play a role in progenitor cell migration during retinal regeneration and/or the timing of photoreceptor regeneration. Currently, CRISPR/Cas9 mutant fish lines for miR-9-1 and miR-9-2 are in the process of being made to allow for additional regeneration studies to be done and to confirm MO knockdown phenotypes.
Details
- Title
- Investigating the role of miR-9 in retinal development and adult regeneration in zebrafish
- Resource Type
- Thesis
- Contributors
- Scott M Taylor (Advisor)Lisa A Waidner (Committee Member)Alexis M Janosik (Committee Member)
- Publisher
- University of West Florida Libraries
- Format
- pdf
- Number of pages
- 55
- Copyright
- Permission granted to the University of West Florida Libraries by the author to digitize and/or display this information for non-profit research and educational purposes. Any reuse of this item in excess of fair use or other copyright exemptions requires the permission of the copyright holder.
- Identifiers
- 99380461596506600
- Academic Unit
- Biology
- Language
- English
- Awarding Institution
- University of West Florida; Bachelor of Science (BS)
- Theses and Dissertations
- Bachelor of Science (BS), University of West Florida