Abstract
The red flour beetle, Tribolium castaneum, is an important model insect and agricultural pest. However, many standard genetic tools are lacking or underdeveloped in this system. Here, we present a set of new reagents to augment existing Tribolium genetic tools. We demonstrate a new GAL4 driver line that employs the promoter of a ribosomal protein gene to drive expression of a UAS responder in the fat body. We also present a novel dual fluorescent reporter that labels cell membranes and nuclei with different fluorophores for the analysis of cellular morphology. This approach also demonstrates the functionality of the viral T2A peptide for bicistronic gene expression in Tribolium. To facilitate classical genetic analysis, we created lines with visible genetic markers by CRISPR-mediated disruption of the yellow and ebony body color loci with a cassette carrying an attP site, enabling future φC31-mediated integration. Together, the reagents presented here will facilitate more robust genetic analysis in Tribolium and serve as a blueprint for the further development of this powerful model’s genetic toolkit.
Highlights
The red flour beetle (Tribolium castaneum), a pest of stored agricultural products, has emerged as a promising system for biological research
Advances in genetic tools have cemented the status of Tribolium as the second model insect of choice behind Drosophila
We address the lack of visible phenotypic markers in Tribolium by using CRISPR to disrupt two genes involved in cuticle pigmentation via homologous recombination with cassettes containing an attP site to facilitate future genomic insertion of DNA of interest using the φC31 integrase
Summary
The red flour beetle (Tribolium castaneum), a pest of stored agricultural products, has emerged as a promising system for biological research. It is a representative of the order Coleoptera, which comprises approximately 40% of known insect species and 25% of all known animals [1]. While Drosophila melanogaster is by far the most popular insect model system, many aspects of its development and physiology are not representative of insects in general, and so findings in Tribolium may be more broadly applicable to insects in many cases. Coleoptera includes significant agricultural pests such as the corn rootworm, Colorado potato beetle, and Asian longhorn beetle, and so using Tribolium as an insect model may lead to advances in pest control.
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