Fig. 2: Genetic characterization of pgSIT.

A The pgSIT cross between double-homozygous gRNA ♂’s harboring both gRNA^βTub#7 and gRNA^myo-fem#1 (termed: gRNA^{βTub + myo-fem}) and homozygous Cas9 ♀’s. The pgSIT cross was initiated reciprocally to generate F₁ transheterozygous progeny carrying either maternal or paternal Cas9 (Supplementary Fig. 4). B Bar graphs comparing the percentage of transheterozygous and heterozygous Cas9 or gRNA progeny to those of WT (data can be found in Supplementary Table 4 and Supplementary Video 4) over at least three (n ≥ 3) biologically independent F₁ progeny groups. C Experimental setup to determine whether prior matings with pgSIT^♂’s suppresses WT female (♀) fertility. WT ♀’s were cohabitated with pgSIT^♂’s for 2, 6, 12, 24, or 48 h then WT ♀’s were transferred to a new cage along with WT ♂’s and mated for an additional 2 days. The ♀’s were then blood-fed and individually transferred to a vial. Eggs were collected and hatched for fertility determination. Following this, nonfertile ♀’s were then placed back into cages along WT ♂’s for another chance to produce progeny. This was repeated for up to five gonotrophic cycles, and the percentage of fertile ♀’s in each group of 50 ♀’s was plotted (Supplementary Table 8). The plot shows the fertility of three biologically independent groups of 50 WT ♀ (n = 3) for each experimental condition. D Flight activity of 24 individual mosquitoes (n = 24) was assessed for 24 h using a vertical Drosophila activity monitoring (DAM) system, which uses an infrared beam to record flight (Supplementary Table 6 and Supplementary Video 5). E To quantify the attractiveness of ♂’s to ♀’s for mating, we used a mating-behavior lure of a tone mimicking ♀ flight. A 10-s 600 Hz sine tone was applied on one side of the cage, and a number of mosquito ♂’s landing on the mesh around a speaker was scored. Heatmaps were generated using Noldus Ethovision XT (Supplementary Table 7 and Supplementary Video 6). The experiment was repeated six times (n = 6) using fresh groups of 30–40 ♂’s. Point plots (C–E) show biological replicates and means ± SDs. A two-sided F test was used to assess the variance equality. Statistical significance of mean differences was estimated using a two-sided Student’s t test with unequal or equal variance. (P ≥ 0.05 ^ns, P < 0.05*, P < 0.01**, and P < 0.001***). Source data are provided as a Source Data File.