CRISPR/Cas9-mediated ebony knockout results in puparium melanism in Spodoptera litura.

Insect body pigmentation and coloration are critical to adaption to the environment. To explore the mechanisms that drive pigmentation, we used the CRISPR/Cas9 genome editing system to target the ebony gene in the non-model insect Spodoptera litura. Ebony is crucial to melanin synthesis in insects. By directly injecting Cas9 mRNA and ebony-specific guide RNAs into S. litura embryos, we successfully induced a typical ebony-deficient phenotype of deep coloration of the puparium and induction of melanin formation during the pupal stage. PCR-based genotype analysis demonstrated that various mutations had occurred at the sites targeted in ebony. Our study clearly demonstrates the function of ebony in the puparium coloration and also provides a potentially useful marker gene for functional studies in S. litura as well as other lepidopteran pests. This article is protected by copyright. All rights reserved.