| Project/Area Number |
22KF0366
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| Project/Area Number (Other) |
22F22768 (2022)
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| Research Category |
Grant-in-Aid for JSPS Fellows
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| Allocation Type | Multi-year Fund (2023)
Single-year Grants (2022) |
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| Section | 外国 |
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| Review Section |
Basic Section 44020:Developmental biology-related
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| Research Institution | Okinawa Institute of Science and Technology Graduate University |
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Principal Investigator |
LAUDET Vincent (2023) 沖縄科学技術大学院大学, 海洋生態進化発生生物学ユニット, 教授 (20898423)
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| Co-Investigator(Kenkyū-buntansha) |
MITCHELL LAURIE 沖縄科学技術大学院大学, 海洋生態進化発生生物学ユニット, 外国人特別研究員
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| Host Researcher |
LAUDET Vincent (2022) 沖縄科学技術大学院大学, 海洋生態進化発生生物学ユニット, 教授 (20898423)
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| Foreign Research Fellow |
MITCHELL LAURIE 沖縄科学技術大学院大学, 海洋生態進化発生生物学ユニット, 外国人特別研究員
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| Project Period (FY) |
2023-03-08 – 2025-03-31
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| Project Status |
Granted (Fiscal Year 2023)
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| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 2024: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2023: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 2022: ¥700,000 (Direct Cost: ¥700,000)
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| Keywords | Pigmentation / Ecology / Coral Reef Fishes / Ontogeny / Cell Death / Color Pattern Change |
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| Outline of Research at the Start |
This project is to investigate the genetic and ecological mechanisms controlling the diversification of the anemonefish white bars. This will allow to understand how variation in fish color patterns arise at the molecular level and its biological significance, therefore providing an explanatory scaffold on pattern diversification.
To achieve this goal:
1.Comparative transcriptomics and breeding experiments to identify genes underlying bar pattern diversity
2.Mutagenesis experiments to directly test gene function
3.Behavioral experiments to test if bar number is used for species recognition
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| Outline of Annual Research Achievements |
The purpose of this project is to investigate the ecological and genetic drivers of pigmentation and color pattern diversity in anemonefishes. To achieve this goal, we have determined the ontogenetic transcriptome of the tomato anemonefish (Amphiprion frenatus) during its juvenile color pattern change and established a link between the external environment and the timing of its pigmentation transition. Moreover, we have demonstrated using cryosection staining techniques the importance of cell death in color pattern change.
To further explore interspecies color pattern differences, we have crossed the (horizontally) striped species A. sandaracinos with the (vertically) barred A. ocellaris to produce a viable first generation of hybrid offspring. The second generation of hybrids will be used to analyze the colour patterns of the hybrids relative to the parent species and identify key genetic loci that correspond to aspects of color pattern variation.
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| Current Status of Research Progress |
Current Status of Research Progress
4: Progress in research has been delayed.
Reason
Major delays in setting-up our husbandry has delayed the establishment of stable breeding fish couples. Moreover, the presence of marine fish egg parasites in our aquarium system has meant that time has been spent on identifying and testing the treatment of infected eggs. This has meant that the final part of this project in using CRISPR/Cas9 genome editing to test the function of specific genes in white bar formation has also been significantly delayed. Our successful treatment of artificially incubated eggs gives us confidence that genome-editing can resume soon, and results will be generated.
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| Strategy for Future Research Activity |
The work is now going in three primary directions:
1° Internal and external processes contributing to ontogenetic bar loss – We have a complete transcriptomic dataset of A. frenatus from metamorphosis to the late-juvenile stage until the complete loss of the white body bar. We are continuing to perform behavioural experiments testing the functional importance of differences in bar number shortly after larval settlement. Moreover, we are performing immunohistochemistry staining assays to confirm that the mode of cell death is apoptosis. A manuscript for this research will be in production shortly thereafter.
2° Gene function in bar formation – We are applying CRISPR/Cas9 genome-editing to perform the targeted knockout of specific genes which are strongly suspected to be involved in white bar formation. One gene of particular interest is fhl2b which when knocked out should produce either the partial or complete loss of white barring.
3° Key genetic loci contributing to color pattern diversity – In collaboration with researchers at Acadamia Sinica Taiwan, we are attempting to raise pairs of the first-generation hybrids (of A. sandaracinos x A. ocellaris) to produce a second generation. This second-generation of hybrids will be photographed and sampled for DNA to perform a quantitative trait loci analysis to identify key genetic factors contributing to aspects of color pattern including bars (vertical) and stripes (horizontal).
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