CRISPR/Cas9 in locusts: Successful establishment of an olfactory deficiency line by targeting the mutagenesis of an odorant receptor co-receptor (Orco) |
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| Institution: | 1. Beijing Institute of Life Science, Chinese Academy of Sciences, Beijing 100101, China;2. State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China;1. Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, China;2. Fujian-Taiwan Joint Centre for Ecological Control of Crop Pests, Fujian Agriculture and Forestry University, Fuzhou 350002, China;3. Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture, Fuzhou 350002, China;4. Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China;5. Department of Microbiology & Molecular Genetics, University of California, Irvine, CA 92697-4025, USA;6. Department of Molecular Biology & Biochemistry, University of California, Irvine, CA 92697-3900, USA;7. Graham Centre, Charles Sturt University, Orange, NSW 2800, Australia;1. Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, Jiangsu, China;2. Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China;1. State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China;2. Beijing Institutes of Life Science, Chinese Academy of Sciences, Beijing 100101, China;1. College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China;2. CSIRO, Black Mountain Laboratories, Canberra, Australian Capital Territory, Australia;1. Department of Entomology, China Agricultural University, Beijing 100193, PR China;2. Bio-tech Research Center, Shandong Academy of Agricultural Sciences, Jinan 250100, PR China;3. Departments of Pharmacology and Neuroscience, University of Texas, Southwestern Medical Center, Dallas, TX 75390, USA |
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| Abstract: | Locusts are important agricultural pests worldwide and regarded as study models for entomology. However, the absence of targeted gene manipulation systems for locusts has restricted their applications for research. Herein, we report the successful use of the CRISPR/Cas9 system to induce a targeted heritable mutagenesis of the migratory locust, Locusta migratoria. The target sequence of gRNA was designed to disrupt the gene encoding the odorant receptor co-receptor (Orco) and examine the roles of the odorant receptor pathway in the locust. Microinjection of the mixture of Cas9-mRNA and Orco-gRNA into the locust eggs resulted in efficient target-gene editing at a rate of 71.7% in G0 animals and achieved a germline efficiency of up to 88.1% in G1 animals. By a crossing strategy, we successfully established stable Orco mutant lines. EAGs and SSRs indicated that the fourth-instar nymphs of the Orco mutants showed severely impaired electrophysiological responses to multiple odors. The Orco mutant locusts lost an attraction response to aggregation pheromones under the crowding conditions. The locomotor activity and body coloration of the Orco mutant locusts did not significantly differ from those of the two other genotypes. This study provides an easy and effective approach by using the CRISPR/Cas9 system for generating loss-of-function mutants for functional genetic studies of locusts and for managing insect pests. |
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| Keywords: | CRISPR/Ca9 Genome editing Olfactory receptor |
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