Development of a Germline Transformation System for the Western Corn Rootworm

The western corn rootworm (WCR) is a major pest of maize and is notorious for rapidly adapting biochemically, behaviorally and developmentally to a variety of control methods. Despite much effort, the genetic basis of WCR adaptation remains a mystery. Transformation-based applications such as transposon tagging and enhancer trapping have facilitated the genetic dissection of model species such as Drosophila melanogaster. Following this model, we are developing a germline transformation system for WCR to gain a greater understanding of how resistance evolves in this economically important insect. We are currently using a piggyBac-based transformation system, known to work in another coleopteran, the red flour beetle, Tribolium castaneum. Unlike Tribolium, WCR lacks eye-color mutants conventionally used to observe transgene expression, therefore our transformation constructs include an alpha-tubulin driven fluorescent marker (EGFP).  This Tribolium-derived promoter is expected to drive transgene expression in every cell. Importantly, the construct is functional in Tribolium, and transient expression has already been observed in injected WCR embryos. We are using the same piggyBac helper, phspBac, used to establish transgenic Tribolium.  Although this construct uses the D. melanogaster heat-shock protein 70 (Dm-hsp70) promoter to drive transposase expression, Dm-hsp70 is known to be functional in a wide range of insects.  Our progress towards developing tools, techniques and protocols for WCR germline transformation will be presented. Establishing transgenic technologies for this beetle is the first step towards bringing a wide-range of transformation–based tools to bear on understanding WCR biology, which can then be extended to other rootworm species.