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. 2021 Apr 13;10:e58791. doi: 10.7554/eLife.58791

Figure 1. Homology-directed modification of Anopheles gambiae midgut loci by CRISPR/Cas9.

Figure 1.

(A) Schematic showing the insertion of the effector construct at the carboxypeptidase (CP), alkaline phosphatase 2 (AP2), and peritrophin 1 (Aper 1) loci. The donor plasmid supplies the effector coding sequence (yellow), which accommodates an artificial intron. The intron harbours a fluorescent marker (either GFP or CFP, green) under the control of the 3xP3 promoter flanked by loxP sites (black dots) and a U6 driven guide-RNA module (red), required for both transgenesis and subsequently for gene drive. The plasmid features regions of homology that drive N- or C-terminal insertion at the start (CP, AP2) or stop codon (Aper1) as well as a 3xP3::DsRed plasmid-backbone marker. The gRNA target sequence (red) including the PAM motif (bold) and the target strand are indicated. (B) Summary of embryo microinjections and the identification of transgenic individuals by fluorescent screening. (C) Adult transgenic mosquitos with fluorescent expression of GFP or CFP in the eyes under the control of the 3xP3 promoter as well as a pupa showing DsRed fluorescence, indicating plasmid-backbone integration (AP2’).