Jacob N Dayton,Tammy T Tran,Elisa Saint-Denis,Erik B Dopman
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Efficient CRISPR/Cas9-mediated genome editing in the European corn borer, Ostrinia nubilalis.
The European corn borer (Ostrinia nubilalis) is an agricultural pest and burgeoning model for research on speciation, seasonal adaptation and insect resistance management. Although previous work in O. nubilalis has identified genes associated with differences in life cycle, reproduction, and resistance to Bt toxins, the general lack of a robust gene-editing protocol for O. nubilalis has been a barrier to functional validation of candidate genes. Here, we demonstrate an efficient and practical methodology for heritable gene mutagenesis in O. nubilalis using the CRISPR/Cas9 genome editing system. Precise loss-of-function (LOF) mutations were generated at two circadian clock genes, period (per) and pigment-dispersing factor receptor (pdfr), and a developmental gene, prothoracicotropic hormone (ptth). Precluding the need for a visible genetic marker, gene-editing efficiency remained high across different single guide RNAs (sgRNA) and germline transmission of mutations to F1 offspring approached 100%. When single or dual sgRNAs were injected at a high concentration, gene-specific phenotypic differences in behaviour and development were identified in F0 mutants. Specifically, F0 gene mutants demonstrated that PER, but not PDFR, is essential for normal timing of eclosion. PTTH F0 mutants were significantly heavier and exhibited a higher incidence of diapause. This work will accelerate future studies of gene function in O. nubilalis and facilitate the development of similar screens in other Lepidopteran and non-model insects.
期刊介绍:
Insect Molecular Biology has been dedicated to providing researchers with the opportunity to publish high quality original research on topics broadly related to insect molecular biology since 1992. IMB is particularly interested in publishing research in insect genomics/genes and proteomics/proteins.
This includes research related to:
• insect gene structure
• control of gene expression
• localisation and function/activity of proteins
• interactions of proteins and ligands/substrates
• effect of mutations on gene/protein function
• evolution of insect genes/genomes, especially where principles relevant to insects in general are established
• molecular population genetics where data are used to identify genes (or regions of genomes) involved in specific adaptations
• gene mapping using molecular tools
• molecular interactions of insects with microorganisms including Wolbachia, symbionts and viruses or other pathogens transmitted by insects
Papers can include large data sets e.g.from micro-array or proteomic experiments or analyses of genome sequences done in silico (subject to the data being placed in the context of hypothesis testing).