Journal article

Publication year: 2021

Pages: 6724-

Journal: International Journal of Molecular Sciences

Volume number: 22

Issue number: 13

Open access status: Gold

DOI Link: https://doi.org/10.3390/ijms22136724

Publisher: MDPI

Abstract: 
The spotted-wing Drosophila (Drosophila suzukii Matsumura) is native to eastern Asia, but has become a global threat to fruit production. In recent years, CRISPR/Cas9 targeting was established in this species allowing for functional genomic and genetic control studies. Here, we report the generation and characterization of Cas9-expressing strains of D. suzukii. Five independent transgenic lines were generated using a piggyBac construct containing the EGFP fluorescent marker gene and the Cas9 gene under the control of the D. melanogaster heat shock protein 70 promoter and 3'UTR. Heat-shock (HS) treated embryos were analyzed by reverse transcriptase PCR, revealing strong heat inducibility of the transgenic Cas9 expression. By injecting gRNA targeting EGFP into one selected line, 50.0% of G(0) flies showed mosaic loss-of-fluorescence phenotype, and 45.5% of G(0) flies produced G(1) mutants without HS. Such somatic and germline mutagenesis rates were increased to 95.4% and 85.7%, respectively, by applying a HS. Parental flies receiving HS resulted in high inheritance of the mutation (92%) in their progeny. Additionally, targeting the endogenous gene yellow led to the lack of pigmentation and male lethality. We discuss the potential use of these efficient and temperature-dependent Cas9-expressing strains for the genetic studies in D. suzukii.

Harvard Citation style: Yan, Y., Kobayashi, Y., Huang, C., Liu, B., Qian, W., Wan, F., et al. (2021) Highly Efficient Temperature Inducible CRISPR-Cas9 Gene Targeting in Drosophila suzukii, International Journal of Molecular Sciences, 22(13), p. 6724. https://doi.org/10.3390/ijms22136724

APA Citation style: Yan, Y., Kobayashi, Y., Huang, C., Liu, B., Qian, W., Wan, F., & Schetelig, M. (2021). Highly Efficient Temperature Inducible CRISPR-Cas9 Gene Targeting in Drosophila suzukii. International Journal of Molecular Sciences. 22(13), 6724. https://doi.org/10.3390/ijms22136724