Ostrinia furnacalis is an important borer on maize. Long-term and large-scale planting of transgenic corn has led O. furnacalis evolving resistance and reducing the control effect. Recently, high levels of resistance to Bt Cry1 toxins have been reported to be genetically linked to the mutation or down-regulation of ABC transporter subfamily G gene ABCG4 in O. furnacalis. In order to further determine the relationship between ABCG4 gene and the resistance to Cry1 toxins in O. furnacalis, the novel CRISPR/Cas9 genome engineering system was utilized to successfully construct ABCG4-KO knockout homozygous strain. Bioassay results indicated that an ABCG4-KO strain had a higher resistance to Cry1 proteins compared with a susceptible strain (ACB-BtS). The result indicates that the ABCG4 gene may act as a receptor of the Bt Cry1 toxin in O. furnacalis. Furthermore, the development time was significantly changed in the early stage ABCG4-KO larvae, and the population parameters were also significantly changed. In summary, our CRISPR/Cas9-mediated genome editing study presents evidence that ABCG4 gene is a functional receptor for Bt Cry1 toxins, laying the foundation for further clarification of the Bt resistance mechanism.

The evolution of target pest population resistance to Bt toxins is the most relevant threat to the sustainability of Bt technology, thus it is necessary to clarify insect resistance mechanisms. Firstly, the resistance level of Asian corn borer was determined by bioassay. After 28 generations selection in the lab, the Cry1Ie-resistant strain (ACB-IeR) developed more than 862-fold resistance to Cry1Ie, and the Cry1F-resistant strain (ACB-FR) developed 961-fold resistance to Cry1F. The results show that long-term exposure to Bt toxins can lead to resistance. Then, we compared the differential expression genes (DEGs) of ACB-FR and ACB-IeR with susceptible strain (ACB-BtS), and analyzed GO function and KEGG pathway through transcriptome sequencing. The comparison showed that in Bt-resistant strains, many genes have a significant down-regulated trend. Several Bt-resistance candidate genes were differentially expressed in both resistant strains. Furthermore, the DEGs were verified by RT-qPCR and showed similar trend. These results provide candidate genes for further research on the Bt resistance mechanism.