Establishment of transformed housefly strains for screening of effective insecticides for controlling medically important pest insects

Research Project

Project/Area Number	17K08816
Research Category	Grant-in-Aid for Scientific Research (C)
Allocation Type	Multi-year Fund
Section	一般
Research Field	Parasitology (including sanitary zoology)
Research Institution	National Institute of Infectious Diseases
Principal Investigator	Kasai Shinji 国立感染症研究所, 昆虫医科学部, 部長 (80332360)
Co-Investigator(Kenkyū-buntansha)	冨田隆史国立感染症研究所, 昆虫医科学部, 主任研究官 (20180169) 駒形修国立感染症研究所, 昆虫医科学部, 主任研究官 (20435712)
Project Period (FY)	2017-04-01 – 2021-03-31
Project Status	Completed (Fiscal Year 2020)
Budget Amount *help	¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000) Fiscal Year 2019: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000) Fiscal Year 2018: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000) Fiscal Year 2017: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
Keywords	殺虫剤抵抗性 / イエバエ / ゲノム編集 / CRISPR/Cas9 / ノックアウト / ノックダウン抵抗性 / kdr / 殺虫剤 / 作用点 / Musca domestica / ナトリウムチャネル / 選択的スプライシング / 創薬
Outline of Final Research Achievements	In order to characterize insecticide resistance genes that are detected in small insects and/or insects that are difficult to rear in the laboratory, we attempted to establish genome editing technology for house flies. Guide RNA designed from a part of white gene were injected into house fly eggs. As a result, we could obtain individuals with white eyes and thus we confirmed that the knockout system we established works in this insect species. Secondly, we attempted another knockout test with the expectation that pyrethroid susceptibility would be recovered by knocking out the resistance mutation that occurred on the alternative exon of the sodium channel gene, which is the target site of pyrethroid insecticides. Despite repeated injection of guide RNA and improvements in methods, we could not confirm any egg hatching and developing to larvae after the injections. It was suggested that the gene targeted in this study is an essential exon region for house fly survival.
Academic Significance and Societal Importance of the Research Achievements	本研究は、イエバエのゲノム編集技術の確立を目的として行われた。イエバエの眼色（赤色）を司るwhite遺伝子のノックアウトを試みた結果、白色眼を有する個体の作出に成功し、インジェクションの系がうまく働いていることを明らかにした。イエバエは古くから飼育法が確立され、モデル昆虫あるいは実験動物として有効活用されてきた。ヒゼンダニやアタマジラミ、サシチョウバエといった比較的飼育が難しかったり、微小で実験が容易ではない生物で確認された遺伝子変異について、イエバエ体内で再現して機能解析を行ったり、難飼育生物の殺虫剤抵抗性系統に有効な殺虫剤の生物検定を効率的に行う上で、大きなステップとなる成果と言える。